Journal of Affect&e Disorders, 24 (1992) 55-62 0 1992 Elsevier Science Publishers B.V. All rights reserved Ol&0327/92/$05.00
55
JAD 00863
elet serotonin and plasma t ogenous depression. Cli erapeutic, and biological correlations Javier Quintana Departments of Neurology and Psychiatry, School of Medicine, University of California at Los Angeles, Los Angeles, CA, USA
(Received 25 July 1991) (Revisioll received 2 October 1991) (Accepted 10 October 1991)
Summary Platelet serotonin (5HT) and plasma tryptophan (free and total) levels were measured in 25 unmedicated depressed patients and in 25 age- and sex-matched healthy controls. The same parameters were determined in the patients after 3 weeks and 2 months of imipramine treatment. Comparisons between patients and control values showed a significant decrease in total plasma tryptophan and platelet 5-HT in unmedicated patients. During treatment, the clinical condition of the patients improved, while plasma tryptophan and platelet 5-HT values normalized after 3 weeks and 2 months, respectively. Clinical status, plasma tryptophan, platelet 5-HT, as well as other biological parameters determined concurrently in the patients, such as platelet monoamine oxidase (MAO), 5-MT uptake, and imipramine binding were compared in search of significant correlations: neither the individual values of any of them nor the magnitude of their changes at any given stage or interval of the study, respectively, were found significantly correlated. These results suggest that a series of 5-HT-related biological parameters are altered in endogenous depression and tend to normalize with imipramine treatment leading to clinical recovery. Within individuals, those parameters are not correlated, suggesting that both the effect of the drug and clinical improvement affect them separately. Key words; Endogenous
depression; Platelet serotonin; Plasma tryptophan;
Introduction The pathophysiology of endogenous depression or at least of some of its subtypes has been
Address for correspondence: Dr. Javier Quintana, UCLA Neuropsychiatric Institute, 760 Westwood Plaza, Los Angeles, CA 90024, USA. Tel.: (213) 825-5528. (213) 825-0247.
Imipramine;
Serotonin
associated with serotonergic function abnormalities in the central nertious system (Coppen, 1967; Maas, 1975). Such abnormalities may result in an incorrect regulati :rr of the synaptic levels of serotonin (5-HT) and therefore in an increase, a decrease, or an abnormal switching between these two states of the levels of that neurotransmitter in certain areas of the brain of depressive subjects. Synaptic 5-HT levels are the result of sev-
eraI regulatory steps throughout the metabolic cyclt: of 5-HT such as synthesis, storage, release, deamination, reuptake, and junctional wash-over. Previously, 1 reported rhe results of studying in parallel some of these processes in blood platelets of depressive patients; these studies allowed a direct comparison of dynamic changes that OCCLX concurrently in the serotonergic neurotransmission system during depression (Quintana, 1988, , anges in the levels of brain and 1989, 1990). C”I platelet monoamine oxidase (MAQ), in the platelet uptake of S-HT, and in the platelet and cerebral binding site for imipramine - the most commonly used antidepressant drug, which binds reversibly to the 5-HT uptake complex - have been extensively documented in depression. Platelets are considered a good peripheral model to study some central neuronal processes (Stahl, 1977; Pletscher and Laubscher, 1980). The synthesis of 5-HT in the brain depends heavily on the availability of the 5-HT precursor 5-H-tryptophan (TRP). This amino acid circulates in blood plasma in two main forms, one free and one bound to plasma albumin. It is thought that, in addition to the plasma levels of fret TRP (Curzon, 1979), the ratio between TRP and some nelJttral amino r;cids (valine, leucine, and isoleutine; NAA) influences the capacity of the former to cross the blocd-brain barrier and become an available source for brain 5-MT synthesis (FernStrom $21.al., 1973; Fernstrom, 1983). Moreover, the proportion of the free and the bound forms of TRP in blood plasma has been found to affect the brain availability of TRP (Curzon, 1979; Fernstrem, 1983). Changes in the levels of plasma TRP, free and total, in Lhe ratio of TRP and neutral amino acids, and/or the levels of brain TRP have been described in dra;Jressed patients (Coppen et al., 1973; DeMycr cc al., 1981; Joseph et al., 1984; Maes et al., 1987; Coppen and Doogan, 1988; Cowen et al., 1989). It has been postulated that those changes could be relatei to the onset of the disease, with abnormal levels of 5-HT Oi serotonergic function, or with the altered neuroendocrine responses to intravenous TRP seen in depressives (Cowen and Charig, 1987; Charney et al., 1984; Heninger et al., 1984; Meltzer et al., 1984a,b). However, some of the results leading to such postulates have been disputed
(Coppen et al., 1973; Riley and Shaw, 1976; Msller et al., 1979; Cowen et al., 1989). A direct consequence of changes in plasma TRP levels is an altered amount of 5-HT in brain and platelets. Abnormal platelet 5-I-IT levels have been found in Jepressives (Wirtz-Justice and Pu hringcr, 1978; Qxt:ukrug, 1979; Sarrias et al., 1987; Guicher ey et al., 1983); similarly, changes in 5-HT levels have been observed in brain tissue from suicides or deceased depressives (review in Coppen and Doogan, 1988). Abnormal levels of some direct metabolites of 5-HT have also been found in the CSF of depressives (Aberg-Wistedt, 1989; Meltzer, 1989; Martensson et al., 1991). Although some of these abnormalities have been associated more to changes in mood-related symptoms such ;?s impulsivity and aggression than to depression itself, they might be a result of S-HT altered levels in the brain. In any case, their finding in depressives is an indication of concurrent brain serotsnin abnormalities and depression. The present study was aimed to (a) provide additional data on levels of free and total plasma TRP, and of platelet 5-HT in depressive patients as compared to values of age- and sex-matched normal controls studied in parallel; and (b) to examine how those levels correlate with the clinical and therapeutic status of the patients, and with other biological parameters measured concurrently in the same subjects. The study was performed in patients before and during imipramine treatment leading to clinica recovery. In addition, it was an attempt to correlate throughout antidepressant treatment and clinical evo!ution changes of platelet 5-HT and plasma TRP levels with those of other biological parameters platelet MAO, 5-HT uptake, and imipramine binding (PIB) - determined concurrently. Subjects and methods
A group of 25 patients and 25 controls was studied (see also Quintana, 1988, 1989, 1990). The patients (age mean 54 f 9.3, range 32-64 yr,rs, 60% women) had been admitted to the Adult Psychiatry Division, Bellvitge Hospital, University of Barcelona, Spain. Diagnoses of ma-
57
jor depressive disorder, endogenous subtype, were established by two psychiatrists according to RDC (Spitzer et al., 1978). After voluntary informed consent was obtained and the experimental criteria (diagnosis, drug intake status - one week drug-free before admission to the hospital -. prediction of treatment suitability) met, the patients became part of the study and were maintained free of medication for at least 6 additional days. The patients were housed in the same unit and received a standard uospital diet. No significant changes in body weight were observed in the patients throughout the study. After the initial drug-free period, a first blood sample was taken and the patients began oral imipramine therapy (125-175 mg/day). Two other blood samples were taken after 3 weeks and 2 months of continuous imipramine administration. Patients were discharged from the hospital between these two last samplings, after a noticeable recovery from depression was verified both by clinical symptoms evaluation and by the Hamilton Rating Scale for Depression (Hamilton, 1967; HRSD). Only I8 patients were available for the third blood sample, taken during an ambulatory control visit. Except for a few patients receiving benzodiazepines for night sedation, and one patient receiving cimetidine for a gastric ulceration, no other drugs were administered throughout the study. The control group consisted of healthy volunteers with no personal or familial history of major psychiatric disorders. They were screened for any drug intake during the 2 weeks prior to the blood sampling. The age and sex distributions were similar in both the patient and the control group. Samples were taken at the same time of the same day in each patient and his/her matching control. Analytical procedures
Blood samples (lo-15 ml) were drawn by anterior cubital venipuncture between 8 : 00 and 9 : 00 a.m. from patients and controls after overnight fasting. For total plasma TRP levels, a heparinized syringe was used during the blood collection. The plasma was then rapidly separated by centrifugation (3000 x g, 10 min!. TRP concentrations were determined by fluorimetry according to the method of Denkla and Dewey (1967),
as modified by Bloxham and Warren (1974), on small aliquots (10 ~1) of plasma after trichloroacetic acid extraction. Free TRP levels were determined on plasma aliquots after slow (1 h) ultrafiltration through protein ceparation filters (Millipore S085047EO) under vacuum. i_-Tryptophan (Sigma) was used as standard. Results are given in pgjml of plasma. To determine the levels of 5-HT, platelets were isolated as described previously (Quintana, 1988, 1989, 1990). In brief, blood samples (5-6 ml) were immediately transferred to silicatecoated tubes containing 1.5 ml of 0.5% EDTA as anticoagulant after their collection. The samples were then successively centrifuged (130 x g, 15 min), the platelet-rich plasma recovered and centrifuged (130 X g, 10 min), the supernatant centrifuged (750 X g, 10 min), resuspended in 1 ml 0.9% NaCl containing 0.1% EDTA, centrifuged again (750 x g, 5 min), and resuspended in bidistilled water at a final approximate concentration of 10” platelets/ml. The final suspension was frozen and thawed twice before the 5-HT analysis, which was carried out within 7 days of the blood extraction. After a short procedure to eliminate the proteins from the samples (Geeraerts et al., 1974), 5-HT was determined according :o the fluorimetric method of Maickel and Miller (1968) with the addition of 0.06% L-cysteine both in the case of the samples and the contra: curve points (5-hydroxytryptamine-creatinine sulfate, Sigma, used as standard). Results are given in pg 5HT/lO” platelets. Statistical comparisons between the mean values in the group of patients and controls were performed using a two-tailed Student’s t-test. For comparisons of platelet 5-!-IT or plasma TRP values with other biological parameters in the same group of patients, Pearson’s product-moment correlation analysis was used; clinical status (HRSD) and biological parameters were cm-elated using Spearman rank-order tests. esults
Total TRP plasma values were found significantly reduced in unmedicated depressives as compared to controls (P < 0.05, Table I). Such decrease disappeared after 3 weeks of treatment
58
TABLE 1 FREE AND TOTAL PLASMA TRYPTOPHAN VALUES IN NORMAL SUBJECTS AND DEPRESSED FORE AND AFTER IMIPRAMINE TREATMENT Group
‘fatal tryptophan
Free tryptophan
Free/total
Controls (12= 25)
9.73 + 0.46
1.82 _t 0.0-J
0.187 + 0.002
Unmedicated depressed patients (n = 25)
8.67 -+_ 0.34 *
1.63 + 0.11
0.19 rto.09
Depressed patients after 3 weeks of imipramine therapy (n = 24)
9.58 f 0.55
1.89kO.Z‘
0.19 kO.02
Depressed patients after 2 months of imipramine therapy (K = 18)
9.41 k 0.67
1.53*0.1
0.17 fO.O1
PATIENTS BE-
tryptophan ratio
Tryptophan values expressed as pg/ml of plasma_tSEM. * P < 0.05 versus corresponding control value.
and was not present either after 2 months of imipramine therapy, when the patients had fully recovered from depression. Free TRP values, however, were found normal in unmedicated depressives and in the same patients after 3 weeks of imipramine. Only a significant decrease in this parameter with respect to controls was observed in the patients after 2 months of treatment, when they had been released from the hospital. The reduction in free T P values was accompanied by a parallel reduction in the ratio of free/total plasma TRP, changes in which had not been seen in previous stages of the study. In unmedicated depressives, in whom total plasma TRP levels were significantly reduced, the levels of free TgP
were not significantly different than in controls, nor was the free/total TRP ratio. Thus, the tendency to reduced free TRP values that was seen in the unmedicated patients was probably a consequence of the reduction in the total TRP plasma levels (Table 1). Table 2 shows the results of the platelet 5-HT level analysis in patients throughout the therapy and in controls. Unmedicated depressed patients showed a significant decrease in platelet 5-HT levels. This decrease tended to normalize with respect to control values only after 2 months of y, when the patients were already clinically sred from depression. However, platelet values after 3 weeks or 2 months of
TABLE 2 PLATELET S’ZROTONIN (5-HT) VALUES IN NORMAL SUBJECT AFTER IMIPRAMINE TREATMENT
5-HT SEM N
AND DEPRESSED
PATIENTS
BEFORE
Controls
Unmedtcated depressed patients
Depressed patients after 3 weeks of imipramine therapy
Depressed patients after 2 months of imipramine therapy
?.27 0.125 25
1.275 * 0.181 25
0.952 * 0.228 24
1.569 0.395 18
Serotonin values expressed as pg 5-HT/109 platelets _+SEM. * P < 0.05 versus corresponding control value.
AND
imipramine therapy did not differ from those measured in the same patients before treatment (Table 2). A multiple analysis of correlation was run between values of plasma TRP, platelet 5-HT, HRSD scores, and other biological parameters studied concurrently in the same group of patients and controls (Quintana, 1988, 1989, 1990). Following the onset of treatment, the paired comparisons between individual values of platelet MAO and platelet 5-HT (- 0.021,’ - 0.414) or plasma free TRP ( - 0.082/0.517) levels showed a trend towards increasing correlation. A similar trend was seen with respect to the correlation between the total and free fractions of plasma TRP (0.075/0.634). Imipramine treatment did not change the average ratio between free and total plasma TRP throughout the study; however, the variance of the population of individual values of this ratio decreased progressively in the course of the three study stages when imipramine was administered (see Table 1). Other than those findings, there was no significant correlation between individual values of any two parameters (including HRSD scores) at any given stage of the study, nor was there between the changes (indtvidual difference scores) of any two parameters during any given period of the study (i.e., pre-treatment to 3 weeks treatment, 3 weeks to 2 months treatment, or pre-treatment to 2 months treatment). iscussion The present results show that platelets, an accepted peripheral model for neurons, show decreased levels of 5-HT during depression; the total plasma levels of TRP, the main precursor of brain 5-HT, are also reduced in affective disorders. These data agree with other reports in the literature: reduced 5-HT levels have been found in platelets of depressives (Sarrias et al., 1987; Guicheney et al., 1988) whereas decreased plasma TRP levels, either total (DeMyer et al., 1981; Joseph et al., 1984; Maes et al., 1987; Bovier et al., 1988; Cowen et al., 1989) or free (Coppen et al., 1973) have been also documented. However, other studies found no changes (Takahashi, 1976) or an increase in platelet 5-HT values in depressives (Wirtz-Justice and Puhringer, 1978; Ox-
enkrug, 1979), or failed to confirm changes in plasma TRP (Riley and Shaw, 1976; Moher et al., 1979; Sarrias et al., 1987). Increases in platelet 5-HT in depressives were reported by Oxenkrug (1979), only for men, in a group of patients in which males predominated. In my study, the majority were female, which might account for the disagreement with that particular report. Other factors such as length of drug-free period or selection of patients could account for the differences in platelet 5-HT levels also. Diagnosis composition of the patient sample could influence the results, although no differences were found in plasma TR? levels between groups of subjects diagnosed as having bipolar disorders and major depression (Bovier et al., 1988). The period of time during which the patients were free of antidepressant therapy could also be blamed for differences in the plasma TRP levels between the present study and others. Tricyclic antidepressants (TCA) are capable of inhibiting the enzyme TRP pyrrolase in the liver, a main source of peripheral TRP metabolism (Badaway and Evans, 1982), thus raising the levels of plasma TRP. However, several studies have shown that total plasma TRP levels are reduced even after TCA treatment or shortly after the end of it (DeMyer et al., 1981; Joseph et al., 1984; Thomas et al., 1987). TCA may also cancel the stimulating effect of cortisol, whose levels are increased in depression, on the same liver enzyme. Thus, differences between some studies in the literature could be due to different drug-free periods before plasma TRP testing, although relatively short. Such periods have been used in studies in which TRP levels were nevertheless found to be reduced (DeMyer et al., 6981; Joseph et al., 1984). Changes in plasma free TRP levels with respect to controls were not observed in the patiems of this study before treatment. Previously, another study reported a decrease in plasma levels of free TRP (Coppen et al., 1973) with no change in total TRP levels in a group of endogenous depressives. Two points deserve mention in order to explain such differences: first, the patients in the study mentioned were all women, whereas in my group 40% of the patients were men. Gender-related differences between clinical status and biological variables have been tenta-
tively associated with different psychoneuroendocrine responses between males and females (Maes et al., 1988). Second, the diet received by the patients could greatly influence the outcome of the plasma TRP analysis (DeMyer et al., 1981; Cowen et al., 1989). In the present study, the patients were maintained on a carefully supervised diet from the moment they were admitted to the hospital. Low values of free and total TRP were still observed after the pretreatment period (6 days) in this study, when the corrective effects of the diet should have taken place according to other studies (DeMyer et al., 1981). ‘II’RP values, however, returned to normal levels after 3 weeks of imipramine treatment. In addition, in the present study, the age and sex of patients and controls were carefully matched, and the results are in agreement with other studies that did the same (Cowen et al., 1989). The effects of imipramine treatment on the average values of the biological parameters investigated in this study are an increase in total plasma TRP levels after 3 weeks, a decrease in free TRP levels and in the ratio free/total TRP after 2 months of therapy, as well as a tendency towards recovery (increase) in the levels of platelet 5-HT. There was a tendency, throughout the treatment, for several biological parameters to become more correlated at the individual level. In addition to the paired relationships between MAO and 5-HT or free TRP levels, the ratio between free and total TRP levels showed more correlation at the individual level after therapy than before in the same population of depressed patients. These findings might indicate or suggest that the continuous therapy with imipramine stabilizes the balance between several interdependent biochemical parameters at the level of serotonergic neurotransmission, either as a result of the prc:bnce of a steady drug concentration in the bload stream, or as a cause and,‘or consequence of the accompanying clinical recovery (noticeable and verified b; the patients’ symptoms and their HRSD scores). Thus, either directly or indirectly (via the action of so&e of those 5-HT neurotransmission components upon others), imipramine affects several steps of 5-I-IT metabolism altered in endogenous depression. The action of the drug is to accelerate clinical
recovery and produce a trend towards normalization of those biochemical para’ reters. Whether the biological changes in unmedicated depressed patients are a cause or a consequence of clinical status, imipramine administration, or recovery cannot be entirely answered with the data available. However, the correlation studies presented here suggest that the presence of irnipramine in plasma affects a series of biochemical parameters while improving clinical status; individual clinical (HRSD) and biological parameters are not correlated at any stage of the study, and changes in those parameters between any two stages of the study are not correlated either. Thus, at least tentatively, it might be concluded that imipramine therapy induces changes in both HRSD scores (clinical status) and biological parameters (MAO, 5-HT, and TRP - total and free - levels), whereas changes in these dependent variables do not seem to be linked by any mutual causative re;&ionship Plasma TRP levels have been associated with mood changes in normal male volunteers (Young et al., 1988). Animal studies have shown that TRP administration increases brain 5-HT synthesis and influences behavior (Coppen and Doogan, 1988). However, the decrease in plasma TRP observed in this and other studies cannot be considered, with the available information, either a cause or a consequence of altered brain serotonin function. Those changes do not account for more than a small percentage of normal baseline plasma TRP levels; furthermore, additional factors such as diet or long-term drug effects could influence both plasma and brain TRP levels. It is known that the ratio between total TRP and NAA competing with TRP for the same bloodbrain carrier is decreased in depressives (DeMyer et al., 1981; Joseph et al., 1984; Maes et al., 1987) but this fact is somehow obscured by the finding of an increase in NAA concentrations as well as of several other plasma amino acids (Mathis et al., 1988). In any case, the finding in the present as well as in other studies (Bovier et al., 1988) that a normalization of plasma TRP levels accompanies clinical recovery in depressives suggests at least that those peripheral levels are somehow, directly or indirectly, connected with functional serotonergic mechanisms in the central nervous system areas supposedly responsible for the onset
61
and maintenance of endogenous depression. Together with other biological disorders related to the same serotonergic systems, the results presented here enlarge the picture of what goes wrong during depression. If nothing else, they strengthen the notion that central serotonergic abnormalities are associated with depressive illness. References Aberg-Wistedt, A. (1989) The antidepressant effects of 5HT uptake inhibitors. Br. J. Psychiatry 8, 32-40. Badaway, A.A.-B. an.i Evans, M. (1982) Inhibition of rat liver tryptophan pyrrolase activity and elevation of brain tryptoohan concentration by acute administration of small doses of antidepressants. Br. J. Pharmacol. 77, 59-67. Bloxham, D.L. and Warren, W.H. (1974) Error in the determination of tryptophan by the method of Denkla and Dewey. A revised procedure. 4nal. Biochem. 60, 621-624. Bovier, P., Widmer, J., Gaillard, J.M. and Tissot, R. (1988) Evolution of red blood cell membrane transport and plasma level of L-tyrosine and L-tryptophan in depressed treated patients according to clinical improvement. Neuropsychobiology 19. 125- 134. Charney. D.S., Heninger, G.R. and Sternberg, D.E. (1984) Serotonin function and the mechanism of action of antidepressant treatment: effect of amitriptyline and desipramine. Arch. Gen. Psychiatry 41, 359-365. Coppen, A.J. (1967) The biochemistry of affective disorder. Br. J. Psychintry 113. 1237-1264. Coppen, A.J. and Doogan, D.P. (1988) Serotonin and its place in the pathogenesis of depression. J. Clin. Psychiatry Suppl. 49, 4-l 1. Coppen, A.J., Eccleston, E.G. and Peet, M. (1973) Total and free tryptophan concentration in the plasma of depressive patients. Lan:et ii, 60-63. Cowen, P.J. and Charig, E.M. (1987) Neuroendocrine responses to intravenous tryptophan in major depression. Arch. Gen. Psychiatry 44, 958-966. Cowen, P.J., Parry-Billings, M. and Newsholme, E.A. (1989) Decreased plasma tryptophan levels in major depression. J. Affect. Disord. 16, 27-31. Curzon, G. (1979) Relationships between plasma, CSF and brain tryptophan. J. Nerrral Trans. Suppl. 15, 81-92. DeMyer. M.K., Shea, P., Hcrldrie, H.C. and Yoshimura, N.Y. (!981) Plasma tr-jptophan and five other amino ac:ds in depressed and normal subjects. Arch. Gen. Psychiatry 38, 642-646. Denkla, W.D. and Dewey, H.K. (1967) The determination of tryptophan in plasma, liver and urine. J. Lab. Clin. Med. 69, 160- 169. Fernstrom, J.D. (1983) Role of precursor availability in control monoamine synthesis in brain. Physiol. Rev. 63. 484546.
Fernstrom, J.D., Larin, F. and Wurtman, R.J. (1973) Correlations between brain tryptophan and plasma neural amino acids foilowing food consumption in rats. Life Sci. 13, 517-524. Geeraerts, F., Schimpfessel, L. and Gokaert, R. (1974) A simple routine method to preserve and determine blood serotonin. Specialia 15, 837. Guicheney, P., Leger. D., Barrat, J., Treveux, R., De Ligmeres, B., Roques, P., Gamier, J.P , Boyer, P., Grenier, J. and Dreux, C. (1988) Platelet serotonin content and plasma tryptophan in peri- and postmenopausal women: variations with plasma cestrogen levels and depressive symptoms. Eur. J. Clin. Invest. 18, 297-304. Hamilton, M. (1960) A rating scale for depression. J. Neurol. Neurosurg. Psychiatry 23, 56-62. Heninger, G.R., Charney, D.S. and Stemberg, D.E. (Z984) Serotonergic function in depression: prolactin response to intravenous tryptophan in depressed patients and healthy subjects. Arch. Gen. Psychiatry 41, 398-402. Joseph, M.S., Brewerton, T.D., Reus, V.I. and Stebbins, G.T. (1984) Plasma L-tryptophan/neural amino acid ratio and dexamethasone suppression in depression. Psychiatr. Res. 11. 185-192. Maas, J.W. (1975) Biogenic amines and depression: biochemical and pharmacological separation of two types of depression. Arch. Gen. Psychiatry 32, 1357 -1361. Maes, M., De Ruyter, M., Hobin, P. and Suy, E. (1987) Relationship between the dexamethasone suppression test and the L-tryptophan/competing amino acids ratio in depression. Psychiatr. Res. 21, 323-335. Maes, M., De Ruyter, M.. Claes, R. and Suy, E. (1988) Sex-related differences in the relationships between selfrated depression and bioiogical markers. J. Affect. Disord. 15, 119-125. Maickel, R.P. and Miller, F.P. (1968) The tluorimetric determination of indolealkylamines in brain and pineal gland. Adv. Pharmacol. 6, 71-77. Martensson, B., Wagner, A., Beck, 0.. Brodin, K., Montero, D. and &berg, M. (1991) Effects of clomtpramine treatment on cerebrospinal fluid monoamine metabolites and platelet 3H-imipramine binding and serotonin uptake and concentrations in major depressive disorder. Acta Psychiatr. Stand. 83, 125-133. Mathis, P., Schmitt, L., Benatia, M., Granier, F., Ghisolfi, J. and Moron, P. (1988) Plasma amino acid disturbances and depression. Encephale. 14, 77-82. Mehzer, H.Y. (1989) Serotonergic dysfunction in depression. Br. J. Psychiatry 8, 25-31. Meltzer, H.Y., Lowy, M., Robertson, A., Goodnick, P. and 1n c,-I,,.._, ? I,l384a) Effect of 5-hyydro~;tryptsphan on serum cortlsol levels in patients with major affective disorders: III. Effect of antidepressants and lithium carbonate. Arch. Gen. Psychiatry 41. 391-397. Meltzer, H.Y., Perline, R., Tricou, B.J., Lowy, M. and Robertson, A. (1984b) Effect of 5-hydroxyttyptophan on serum cortisol levels in patients with major affective disorders: I. Enhanced response in depression and mania. Arch. Gen. Psychiatry 41, 3ri6-374.
62 Moller, S.E., Kirk, L. and Honore, P. (1979) Free and total plasma rrvptophan in endogenous depression. J. Affect. Disord. 1, 69-76. Oxenkrug, G.F. (197o) The content and uptake of 5HT by blood platelets in depressive patients. J. Neural Trans. 45, 285-289. Pietscher, A. and Laubscher, A. (1980) Blood platelets as models for neurons: uses and limitations. J. Neura: Trans. Suppl. 16, ‘-16. Quintane, J. (1988) Piatelet MAO deamination of serotonin in depressed patients. Changes after imipramine treatment and clinical correlations. Biol. Psychiatry 23. 44-52. Quintana, J. (1989) Platelet serotonin uptake dynamic changes in depression: effects of long-term imipramine treatment and clinical recovery. J. Affect. Disord. 16, 233-242. Quintana, J. (1990) Platelet imipramine binding in endogenous depressed patients and controls: relationship to platelet MAO and SHT uptake during successful imipramine treal.,lent. Psychiatr. Res. 33, 229-242. Riley, C.J. and Shaw, D.M. (1976) Total and nonbound tryptophan in unipolar illness. Lancet ii, 1249. Sarrias, M.J., Artigas, F., Martinez, E.. Gelpi, E., Alvarez, E.,
Udina, C. and Casas, M. (1987) Decreased plasma serotonin in melancholic patients: a study with clomipramine. Biol. Psychiatry 22, 1429-1438. Spitzer, R.L., Endicott, J. and Robins E. (1978) Research Diagnostic Criteria: rationale and reliability. Arch. Gen. Psychiatry 35, 773-782. Stahl, S.M. (19773 The human platelet: a diagnostic and research tool for the study of biogenic amines in psychiatric and neurologic disorders. Arch. Gen. Psych,?try 34, 509-5 16. Takahashi, S. (1976) Reduction of blood platelet serctonin levels in manic and depressive patients. Foha Psych&r. Neurol. Jap. 30, 476. Thomas, R., Shaw, D.M. and Tidmarsh, SF. (1987) Thy effect of tricyclic medication on total plasma tryptophan. J. Clin. Psychopharmacol. 7, 205-206. Wirtz-Justice, A. and Puhringer, W. (1978) Seasonal incidence of an altered diurnal :hythm of platelet serotonin in unipo lar depression. J. Neural Trans. 42, 45-53. Young, S.N., Pihl. R.O. and Ervon, F.R. (198%)The effect of altered tryptophan levels on mood and behavior in normal human males. Chin. Neuropharmacol. Suppl. 1, 207-215.
55
JAD 00863
elet serotonin and plasma t ogenous depression. Cli erapeutic, and biological correlations Javier Quintana Departments of Neurology and Psychiatry, School of Medicine, University of California at Los Angeles, Los Angeles, CA, USA
(Received 25 July 1991) (Revisioll received 2 October 1991) (Accepted 10 October 1991)
Summary Platelet serotonin (5HT) and plasma tryptophan (free and total) levels were measured in 25 unmedicated depressed patients and in 25 age- and sex-matched healthy controls. The same parameters were determined in the patients after 3 weeks and 2 months of imipramine treatment. Comparisons between patients and control values showed a significant decrease in total plasma tryptophan and platelet 5-HT in unmedicated patients. During treatment, the clinical condition of the patients improved, while plasma tryptophan and platelet 5-HT values normalized after 3 weeks and 2 months, respectively. Clinical status, plasma tryptophan, platelet 5-HT, as well as other biological parameters determined concurrently in the patients, such as platelet monoamine oxidase (MAO), 5-MT uptake, and imipramine binding were compared in search of significant correlations: neither the individual values of any of them nor the magnitude of their changes at any given stage or interval of the study, respectively, were found significantly correlated. These results suggest that a series of 5-HT-related biological parameters are altered in endogenous depression and tend to normalize with imipramine treatment leading to clinical recovery. Within individuals, those parameters are not correlated, suggesting that both the effect of the drug and clinical improvement affect them separately. Key words; Endogenous
depression; Platelet serotonin; Plasma tryptophan;
Introduction The pathophysiology of endogenous depression or at least of some of its subtypes has been
Address for correspondence: Dr. Javier Quintana, UCLA Neuropsychiatric Institute, 760 Westwood Plaza, Los Angeles, CA 90024, USA. Tel.: (213) 825-5528. (213) 825-0247.
Imipramine;
Serotonin
associated with serotonergic function abnormalities in the central nertious system (Coppen, 1967; Maas, 1975). Such abnormalities may result in an incorrect regulati :rr of the synaptic levels of serotonin (5-HT) and therefore in an increase, a decrease, or an abnormal switching between these two states of the levels of that neurotransmitter in certain areas of the brain of depressive subjects. Synaptic 5-HT levels are the result of sev-
eraI regulatory steps throughout the metabolic cyclt: of 5-HT such as synthesis, storage, release, deamination, reuptake, and junctional wash-over. Previously, 1 reported rhe results of studying in parallel some of these processes in blood platelets of depressive patients; these studies allowed a direct comparison of dynamic changes that OCCLX concurrently in the serotonergic neurotransmission system during depression (Quintana, 1988, , anges in the levels of brain and 1989, 1990). C”I platelet monoamine oxidase (MAQ), in the platelet uptake of S-HT, and in the platelet and cerebral binding site for imipramine - the most commonly used antidepressant drug, which binds reversibly to the 5-HT uptake complex - have been extensively documented in depression. Platelets are considered a good peripheral model to study some central neuronal processes (Stahl, 1977; Pletscher and Laubscher, 1980). The synthesis of 5-HT in the brain depends heavily on the availability of the 5-HT precursor 5-H-tryptophan (TRP). This amino acid circulates in blood plasma in two main forms, one free and one bound to plasma albumin. It is thought that, in addition to the plasma levels of fret TRP (Curzon, 1979), the ratio between TRP and some nelJttral amino r;cids (valine, leucine, and isoleutine; NAA) influences the capacity of the former to cross the blocd-brain barrier and become an available source for brain 5-MT synthesis (FernStrom $21.al., 1973; Fernstrom, 1983). Moreover, the proportion of the free and the bound forms of TRP in blood plasma has been found to affect the brain availability of TRP (Curzon, 1979; Fernstrem, 1983). Changes in the levels of plasma TRP, free and total, in Lhe ratio of TRP and neutral amino acids, and/or the levels of brain TRP have been described in dra;Jressed patients (Coppen et al., 1973; DeMycr cc al., 1981; Joseph et al., 1984; Maes et al., 1987; Coppen and Doogan, 1988; Cowen et al., 1989). It has been postulated that those changes could be relatei to the onset of the disease, with abnormal levels of 5-HT Oi serotonergic function, or with the altered neuroendocrine responses to intravenous TRP seen in depressives (Cowen and Charig, 1987; Charney et al., 1984; Heninger et al., 1984; Meltzer et al., 1984a,b). However, some of the results leading to such postulates have been disputed
(Coppen et al., 1973; Riley and Shaw, 1976; Msller et al., 1979; Cowen et al., 1989). A direct consequence of changes in plasma TRP levels is an altered amount of 5-HT in brain and platelets. Abnormal platelet 5-I-IT levels have been found in Jepressives (Wirtz-Justice and Pu hringcr, 1978; Qxt:ukrug, 1979; Sarrias et al., 1987; Guicher ey et al., 1983); similarly, changes in 5-HT levels have been observed in brain tissue from suicides or deceased depressives (review in Coppen and Doogan, 1988). Abnormal levels of some direct metabolites of 5-HT have also been found in the CSF of depressives (Aberg-Wistedt, 1989; Meltzer, 1989; Martensson et al., 1991). Although some of these abnormalities have been associated more to changes in mood-related symptoms such ;?s impulsivity and aggression than to depression itself, they might be a result of S-HT altered levels in the brain. In any case, their finding in depressives is an indication of concurrent brain serotsnin abnormalities and depression. The present study was aimed to (a) provide additional data on levels of free and total plasma TRP, and of platelet 5-HT in depressive patients as compared to values of age- and sex-matched normal controls studied in parallel; and (b) to examine how those levels correlate with the clinical and therapeutic status of the patients, and with other biological parameters measured concurrently in the same subjects. The study was performed in patients before and during imipramine treatment leading to clinica recovery. In addition, it was an attempt to correlate throughout antidepressant treatment and clinical evo!ution changes of platelet 5-HT and plasma TRP levels with those of other biological parameters platelet MAO, 5-HT uptake, and imipramine binding (PIB) - determined concurrently. Subjects and methods
A group of 25 patients and 25 controls was studied (see also Quintana, 1988, 1989, 1990). The patients (age mean 54 f 9.3, range 32-64 yr,rs, 60% women) had been admitted to the Adult Psychiatry Division, Bellvitge Hospital, University of Barcelona, Spain. Diagnoses of ma-
57
jor depressive disorder, endogenous subtype, were established by two psychiatrists according to RDC (Spitzer et al., 1978). After voluntary informed consent was obtained and the experimental criteria (diagnosis, drug intake status - one week drug-free before admission to the hospital -. prediction of treatment suitability) met, the patients became part of the study and were maintained free of medication for at least 6 additional days. The patients were housed in the same unit and received a standard uospital diet. No significant changes in body weight were observed in the patients throughout the study. After the initial drug-free period, a first blood sample was taken and the patients began oral imipramine therapy (125-175 mg/day). Two other blood samples were taken after 3 weeks and 2 months of continuous imipramine administration. Patients were discharged from the hospital between these two last samplings, after a noticeable recovery from depression was verified both by clinical symptoms evaluation and by the Hamilton Rating Scale for Depression (Hamilton, 1967; HRSD). Only I8 patients were available for the third blood sample, taken during an ambulatory control visit. Except for a few patients receiving benzodiazepines for night sedation, and one patient receiving cimetidine for a gastric ulceration, no other drugs were administered throughout the study. The control group consisted of healthy volunteers with no personal or familial history of major psychiatric disorders. They were screened for any drug intake during the 2 weeks prior to the blood sampling. The age and sex distributions were similar in both the patient and the control group. Samples were taken at the same time of the same day in each patient and his/her matching control. Analytical procedures
Blood samples (lo-15 ml) were drawn by anterior cubital venipuncture between 8 : 00 and 9 : 00 a.m. from patients and controls after overnight fasting. For total plasma TRP levels, a heparinized syringe was used during the blood collection. The plasma was then rapidly separated by centrifugation (3000 x g, 10 min!. TRP concentrations were determined by fluorimetry according to the method of Denkla and Dewey (1967),
as modified by Bloxham and Warren (1974), on small aliquots (10 ~1) of plasma after trichloroacetic acid extraction. Free TRP levels were determined on plasma aliquots after slow (1 h) ultrafiltration through protein ceparation filters (Millipore S085047EO) under vacuum. i_-Tryptophan (Sigma) was used as standard. Results are given in pgjml of plasma. To determine the levels of 5-HT, platelets were isolated as described previously (Quintana, 1988, 1989, 1990). In brief, blood samples (5-6 ml) were immediately transferred to silicatecoated tubes containing 1.5 ml of 0.5% EDTA as anticoagulant after their collection. The samples were then successively centrifuged (130 x g, 15 min), the platelet-rich plasma recovered and centrifuged (130 X g, 10 min), the supernatant centrifuged (750 X g, 10 min), resuspended in 1 ml 0.9% NaCl containing 0.1% EDTA, centrifuged again (750 x g, 5 min), and resuspended in bidistilled water at a final approximate concentration of 10” platelets/ml. The final suspension was frozen and thawed twice before the 5-HT analysis, which was carried out within 7 days of the blood extraction. After a short procedure to eliminate the proteins from the samples (Geeraerts et al., 1974), 5-HT was determined according :o the fluorimetric method of Maickel and Miller (1968) with the addition of 0.06% L-cysteine both in the case of the samples and the contra: curve points (5-hydroxytryptamine-creatinine sulfate, Sigma, used as standard). Results are given in pg 5HT/lO” platelets. Statistical comparisons between the mean values in the group of patients and controls were performed using a two-tailed Student’s t-test. For comparisons of platelet 5-!-IT or plasma TRP values with other biological parameters in the same group of patients, Pearson’s product-moment correlation analysis was used; clinical status (HRSD) and biological parameters were cm-elated using Spearman rank-order tests. esults
Total TRP plasma values were found significantly reduced in unmedicated depressives as compared to controls (P < 0.05, Table I). Such decrease disappeared after 3 weeks of treatment
58
TABLE 1 FREE AND TOTAL PLASMA TRYPTOPHAN VALUES IN NORMAL SUBJECTS AND DEPRESSED FORE AND AFTER IMIPRAMINE TREATMENT Group
‘fatal tryptophan
Free tryptophan
Free/total
Controls (12= 25)
9.73 + 0.46
1.82 _t 0.0-J
0.187 + 0.002
Unmedicated depressed patients (n = 25)
8.67 -+_ 0.34 *
1.63 + 0.11
0.19 rto.09
Depressed patients after 3 weeks of imipramine therapy (n = 24)
9.58 f 0.55
1.89kO.Z‘
0.19 kO.02
Depressed patients after 2 months of imipramine therapy (K = 18)
9.41 k 0.67
1.53*0.1
0.17 fO.O1
PATIENTS BE-
tryptophan ratio
Tryptophan values expressed as pg/ml of plasma_tSEM. * P < 0.05 versus corresponding control value.
and was not present either after 2 months of imipramine therapy, when the patients had fully recovered from depression. Free TRP values, however, were found normal in unmedicated depressives and in the same patients after 3 weeks of imipramine. Only a significant decrease in this parameter with respect to controls was observed in the patients after 2 months of treatment, when they had been released from the hospital. The reduction in free T P values was accompanied by a parallel reduction in the ratio of free/total plasma TRP, changes in which had not been seen in previous stages of the study. In unmedicated depressives, in whom total plasma TRP levels were significantly reduced, the levels of free TgP
were not significantly different than in controls, nor was the free/total TRP ratio. Thus, the tendency to reduced free TRP values that was seen in the unmedicated patients was probably a consequence of the reduction in the total TRP plasma levels (Table 1). Table 2 shows the results of the platelet 5-HT level analysis in patients throughout the therapy and in controls. Unmedicated depressed patients showed a significant decrease in platelet 5-HT levels. This decrease tended to normalize with respect to control values only after 2 months of y, when the patients were already clinically sred from depression. However, platelet values after 3 weeks or 2 months of
TABLE 2 PLATELET S’ZROTONIN (5-HT) VALUES IN NORMAL SUBJECT AFTER IMIPRAMINE TREATMENT
5-HT SEM N
AND DEPRESSED
PATIENTS
BEFORE
Controls
Unmedtcated depressed patients
Depressed patients after 3 weeks of imipramine therapy
Depressed patients after 2 months of imipramine therapy
?.27 0.125 25
1.275 * 0.181 25
0.952 * 0.228 24
1.569 0.395 18
Serotonin values expressed as pg 5-HT/109 platelets _+SEM. * P < 0.05 versus corresponding control value.
AND
imipramine therapy did not differ from those measured in the same patients before treatment (Table 2). A multiple analysis of correlation was run between values of plasma TRP, platelet 5-HT, HRSD scores, and other biological parameters studied concurrently in the same group of patients and controls (Quintana, 1988, 1989, 1990). Following the onset of treatment, the paired comparisons between individual values of platelet MAO and platelet 5-HT (- 0.021,’ - 0.414) or plasma free TRP ( - 0.082/0.517) levels showed a trend towards increasing correlation. A similar trend was seen with respect to the correlation between the total and free fractions of plasma TRP (0.075/0.634). Imipramine treatment did not change the average ratio between free and total plasma TRP throughout the study; however, the variance of the population of individual values of this ratio decreased progressively in the course of the three study stages when imipramine was administered (see Table 1). Other than those findings, there was no significant correlation between individual values of any two parameters (including HRSD scores) at any given stage of the study, nor was there between the changes (indtvidual difference scores) of any two parameters during any given period of the study (i.e., pre-treatment to 3 weeks treatment, 3 weeks to 2 months treatment, or pre-treatment to 2 months treatment). iscussion The present results show that platelets, an accepted peripheral model for neurons, show decreased levels of 5-HT during depression; the total plasma levels of TRP, the main precursor of brain 5-HT, are also reduced in affective disorders. These data agree with other reports in the literature: reduced 5-HT levels have been found in platelets of depressives (Sarrias et al., 1987; Guicheney et al., 1988) whereas decreased plasma TRP levels, either total (DeMyer et al., 1981; Joseph et al., 1984; Maes et al., 1987; Bovier et al., 1988; Cowen et al., 1989) or free (Coppen et al., 1973) have been also documented. However, other studies found no changes (Takahashi, 1976) or an increase in platelet 5-HT values in depressives (Wirtz-Justice and Puhringer, 1978; Ox-
enkrug, 1979), or failed to confirm changes in plasma TRP (Riley and Shaw, 1976; Moher et al., 1979; Sarrias et al., 1987). Increases in platelet 5-HT in depressives were reported by Oxenkrug (1979), only for men, in a group of patients in which males predominated. In my study, the majority were female, which might account for the disagreement with that particular report. Other factors such as length of drug-free period or selection of patients could account for the differences in platelet 5-HT levels also. Diagnosis composition of the patient sample could influence the results, although no differences were found in plasma TR? levels between groups of subjects diagnosed as having bipolar disorders and major depression (Bovier et al., 1988). The period of time during which the patients were free of antidepressant therapy could also be blamed for differences in the plasma TRP levels between the present study and others. Tricyclic antidepressants (TCA) are capable of inhibiting the enzyme TRP pyrrolase in the liver, a main source of peripheral TRP metabolism (Badaway and Evans, 1982), thus raising the levels of plasma TRP. However, several studies have shown that total plasma TRP levels are reduced even after TCA treatment or shortly after the end of it (DeMyer et al., 1981; Joseph et al., 1984; Thomas et al., 1987). TCA may also cancel the stimulating effect of cortisol, whose levels are increased in depression, on the same liver enzyme. Thus, differences between some studies in the literature could be due to different drug-free periods before plasma TRP testing, although relatively short. Such periods have been used in studies in which TRP levels were nevertheless found to be reduced (DeMyer et al., 6981; Joseph et al., 1984). Changes in plasma free TRP levels with respect to controls were not observed in the patiems of this study before treatment. Previously, another study reported a decrease in plasma levels of free TRP (Coppen et al., 1973) with no change in total TRP levels in a group of endogenous depressives. Two points deserve mention in order to explain such differences: first, the patients in the study mentioned were all women, whereas in my group 40% of the patients were men. Gender-related differences between clinical status and biological variables have been tenta-
tively associated with different psychoneuroendocrine responses between males and females (Maes et al., 1988). Second, the diet received by the patients could greatly influence the outcome of the plasma TRP analysis (DeMyer et al., 1981; Cowen et al., 1989). In the present study, the patients were maintained on a carefully supervised diet from the moment they were admitted to the hospital. Low values of free and total TRP were still observed after the pretreatment period (6 days) in this study, when the corrective effects of the diet should have taken place according to other studies (DeMyer et al., 1981). ‘II’RP values, however, returned to normal levels after 3 weeks of imipramine treatment. In addition, in the present study, the age and sex of patients and controls were carefully matched, and the results are in agreement with other studies that did the same (Cowen et al., 1989). The effects of imipramine treatment on the average values of the biological parameters investigated in this study are an increase in total plasma TRP levels after 3 weeks, a decrease in free TRP levels and in the ratio free/total TRP after 2 months of therapy, as well as a tendency towards recovery (increase) in the levels of platelet 5-HT. There was a tendency, throughout the treatment, for several biological parameters to become more correlated at the individual level. In addition to the paired relationships between MAO and 5-HT or free TRP levels, the ratio between free and total TRP levels showed more correlation at the individual level after therapy than before in the same population of depressed patients. These findings might indicate or suggest that the continuous therapy with imipramine stabilizes the balance between several interdependent biochemical parameters at the level of serotonergic neurotransmission, either as a result of the prc:bnce of a steady drug concentration in the bload stream, or as a cause and,‘or consequence of the accompanying clinical recovery (noticeable and verified b; the patients’ symptoms and their HRSD scores). Thus, either directly or indirectly (via the action of so&e of those 5-HT neurotransmission components upon others), imipramine affects several steps of 5-I-IT metabolism altered in endogenous depression. The action of the drug is to accelerate clinical
recovery and produce a trend towards normalization of those biochemical para’ reters. Whether the biological changes in unmedicated depressed patients are a cause or a consequence of clinical status, imipramine administration, or recovery cannot be entirely answered with the data available. However, the correlation studies presented here suggest that the presence of irnipramine in plasma affects a series of biochemical parameters while improving clinical status; individual clinical (HRSD) and biological parameters are not correlated at any stage of the study, and changes in those parameters between any two stages of the study are not correlated either. Thus, at least tentatively, it might be concluded that imipramine therapy induces changes in both HRSD scores (clinical status) and biological parameters (MAO, 5-HT, and TRP - total and free - levels), whereas changes in these dependent variables do not seem to be linked by any mutual causative re;&ionship Plasma TRP levels have been associated with mood changes in normal male volunteers (Young et al., 1988). Animal studies have shown that TRP administration increases brain 5-HT synthesis and influences behavior (Coppen and Doogan, 1988). However, the decrease in plasma TRP observed in this and other studies cannot be considered, with the available information, either a cause or a consequence of altered brain serotonin function. Those changes do not account for more than a small percentage of normal baseline plasma TRP levels; furthermore, additional factors such as diet or long-term drug effects could influence both plasma and brain TRP levels. It is known that the ratio between total TRP and NAA competing with TRP for the same bloodbrain carrier is decreased in depressives (DeMyer et al., 1981; Joseph et al., 1984; Maes et al., 1987) but this fact is somehow obscured by the finding of an increase in NAA concentrations as well as of several other plasma amino acids (Mathis et al., 1988). In any case, the finding in the present as well as in other studies (Bovier et al., 1988) that a normalization of plasma TRP levels accompanies clinical recovery in depressives suggests at least that those peripheral levels are somehow, directly or indirectly, connected with functional serotonergic mechanisms in the central nervous system areas supposedly responsible for the onset
61
and maintenance of endogenous depression. Together with other biological disorders related to the same serotonergic systems, the results presented here enlarge the picture of what goes wrong during depression. If nothing else, they strengthen the notion that central serotonergic abnormalities are associated with depressive illness. References Aberg-Wistedt, A. (1989) The antidepressant effects of 5HT uptake inhibitors. Br. J. Psychiatry 8, 32-40. Badaway, A.A.-B. an.i Evans, M. (1982) Inhibition of rat liver tryptophan pyrrolase activity and elevation of brain tryptoohan concentration by acute administration of small doses of antidepressants. Br. J. Pharmacol. 77, 59-67. Bloxham, D.L. and Warren, W.H. (1974) Error in the determination of tryptophan by the method of Denkla and Dewey. A revised procedure. 4nal. Biochem. 60, 621-624. Bovier, P., Widmer, J., Gaillard, J.M. and Tissot, R. (1988) Evolution of red blood cell membrane transport and plasma level of L-tyrosine and L-tryptophan in depressed treated patients according to clinical improvement. Neuropsychobiology 19. 125- 134. Charney. D.S., Heninger, G.R. and Sternberg, D.E. (1984) Serotonin function and the mechanism of action of antidepressant treatment: effect of amitriptyline and desipramine. Arch. Gen. Psychiatry 41, 359-365. Coppen, A.J. (1967) The biochemistry of affective disorder. Br. J. Psychintry 113. 1237-1264. Coppen, A.J. and Doogan, D.P. (1988) Serotonin and its place in the pathogenesis of depression. J. Clin. Psychiatry Suppl. 49, 4-l 1. Coppen, A.J., Eccleston, E.G. and Peet, M. (1973) Total and free tryptophan concentration in the plasma of depressive patients. Lan:et ii, 60-63. Cowen, P.J. and Charig, E.M. (1987) Neuroendocrine responses to intravenous tryptophan in major depression. Arch. Gen. Psychiatry 44, 958-966. Cowen, P.J., Parry-Billings, M. and Newsholme, E.A. (1989) Decreased plasma tryptophan levels in major depression. J. Affect. Disord. 16, 27-31. Curzon, G. (1979) Relationships between plasma, CSF and brain tryptophan. J. Nerrral Trans. Suppl. 15, 81-92. DeMyer. M.K., Shea, P., Hcrldrie, H.C. and Yoshimura, N.Y. (!981) Plasma tr-jptophan and five other amino ac:ds in depressed and normal subjects. Arch. Gen. Psychiatry 38, 642-646. Denkla, W.D. and Dewey, H.K. (1967) The determination of tryptophan in plasma, liver and urine. J. Lab. Clin. Med. 69, 160- 169. Fernstrom, J.D. (1983) Role of precursor availability in control monoamine synthesis in brain. Physiol. Rev. 63. 484546.
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62 Moller, S.E., Kirk, L. and Honore, P. (1979) Free and total plasma rrvptophan in endogenous depression. J. Affect. Disord. 1, 69-76. Oxenkrug, G.F. (197o) The content and uptake of 5HT by blood platelets in depressive patients. J. Neural Trans. 45, 285-289. Pietscher, A. and Laubscher, A. (1980) Blood platelets as models for neurons: uses and limitations. J. Neura: Trans. Suppl. 16, ‘-16. Quintane, J. (1988) Piatelet MAO deamination of serotonin in depressed patients. Changes after imipramine treatment and clinical correlations. Biol. Psychiatry 23. 44-52. Quintana, J. (1989) Platelet serotonin uptake dynamic changes in depression: effects of long-term imipramine treatment and clinical recovery. J. Affect. Disord. 16, 233-242. Quintana, J. (1990) Platelet imipramine binding in endogenous depressed patients and controls: relationship to platelet MAO and SHT uptake during successful imipramine treal.,lent. Psychiatr. Res. 33, 229-242. Riley, C.J. and Shaw, D.M. (1976) Total and nonbound tryptophan in unipolar illness. Lancet ii, 1249. Sarrias, M.J., Artigas, F., Martinez, E.. Gelpi, E., Alvarez, E.,
Udina, C. and Casas, M. (1987) Decreased plasma serotonin in melancholic patients: a study with clomipramine. Biol. Psychiatry 22, 1429-1438. Spitzer, R.L., Endicott, J. and Robins E. (1978) Research Diagnostic Criteria: rationale and reliability. Arch. Gen. Psychiatry 35, 773-782. Stahl, S.M. (19773 The human platelet: a diagnostic and research tool for the study of biogenic amines in psychiatric and neurologic disorders. Arch. Gen. Psych,?try 34, 509-5 16. Takahashi, S. (1976) Reduction of blood platelet serctonin levels in manic and depressive patients. Foha Psych&r. Neurol. Jap. 30, 476. Thomas, R., Shaw, D.M. and Tidmarsh, SF. (1987) Thy effect of tricyclic medication on total plasma tryptophan. J. Clin. Psychopharmacol. 7, 205-206. Wirtz-Justice, A. and Puhringer, W. (1978) Seasonal incidence of an altered diurnal :hythm of platelet serotonin in unipo lar depression. J. Neural Trans. 42, 45-53. Young, S.N., Pihl. R.O. and Ervon, F.R. (198%)The effect of altered tryptophan levels on mood and behavior in normal human males. Chin. Neuropharmacol. Suppl. 1, 207-215.
