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Changes in platelet 3H-imipramine binding in depressed patients receiving electroconvulsive therapy
BIOL PSYCTUATRY 1980;24:469412
469
Changes in Platelet 3H-Imipramine Binding in Depressed Patients Receiving Electroconvulsive Therapy Mario Maj, Pasquale Mastronardi, Antonia Cerreta, Michele Romano, Basilio Mazzarella, and Dargut Kemali
Introduction A significant decrease in the density of platelet 3H-imipramine binding sites (B,) in untreated depressed patients as compared with healthy controls has been reported by several authors (Briley et al. 1980; Raisman et al. 1981; Wagner et al. 1985), although not confirmed by some others (Mellerup et al. 1982; Kanof et al. 1987). B, values have been found to remain low during partial clinical improvement following treatment with tricyclic antidepressants (Briley et al. 1980; Suranyi-Cadotte et al. 1985). but to increase to a normal range after 8-12 weeks of treatment, when complete symptomatic recovery is achieved (Suranyi-Cadotte et al. 1985). Similar results have been obtained by Langer et al. (1986) in depressed patients receiving electroconvulsive therapy (ECT): B,, values were still significantly lower than those of normal controls after 610 sessions of ECT, when clinical improvement was only partial, whereas they were found to be in the normal range after 12-l 8 months, at a time when patients were
From the Depltmcnt of Medical Psychology and Psycltiatty. Fmt Medical School. Uttivcnity of Nqks. Italy (MM., A.C., D.K.); theSewnd~of Araabcsidogy, Sccottd Medial S&ml. Uttivcnity of Nqh. Italy (P.M.. B.M.); atuI the Cam di Cm Via Chiant@. Nocen. hktno, Italy (MR.). AdmtsstqintmqtatstoPmfmuxhihohhj,Catt&adiIgiette Menule, Prima pOliclinica Univaritatio. Piazza Miraglia 2, I80138 Nqoli, Italy. Received Jttnc 30, 1987; revised !kphtbu 16. 1987.
0 1988 Society of Biological Psychiatry
euthymic. In this study, however, the 6 patients in whom platelet 3H-imipramine binding was determined after recovery from depression were on treatment with antidepressant drugs (4 of them also with lithium) at the time of this assessment, so that a possible effect of this additional treatment on B, cannot be excluded. Moreover, as the assessment was made several months after clinical improvement, it was impossible to explore whether or not the more rapid clinical effect of ECT in comparison with tricyclic antidepressants (Fawcett and Scheftner 1986) is paralleled by a more rapid normalization of platelet ‘H-imipramine binding. This last issue was addressed in the present investigation in which platelet 3H-imipramine binding was studied in a sample of patients with major depression at baseline, after 4 and 8 sessions of ECT, and immediately after recovery from the depressive episode.
Methods Nineteen inpatients (14 men and 5 women, age range 18-59 years, mean + SD 32.8 + 10.8), fulfilling DSM-III criteria for a diagnosis of major depressive episode, entered the study. They scored at least 16 (mean 25.7 & 4.2 SD) on the first 17 items of the 2 l-item version of the Hamilton Rating Scale for Depression @IRS-D) (Hamilton 1960) and had been free from psychotropic drugs (except low-dose benzodiaze-
470
BIOL PSYCHIATRY i9aa;24:469-472
pines) for at least 12 days. The control population included 19 subjects without personal or family history of psychiatric disorders (14 men and 5 women, age range 20-59 years, mean t SD 34.1 ? 12.0), who had not received psychotropic drugs in the previous month. Both patients and controls were physically healthy. Patients were treated with ECT because of the severity of depression or the unfavorable response to tricyclic antidepressants in previous episodes. Bilateral ECT was performed after premeditation with atropine sulfate, 1 mg im, thiopental sodium, 4 mg/kg iv, and succinylcholine chloride 0.7 mg/kg iv. Platelet 3H-imipramine binding was determined in patients and controls at baseline and in patients after 14 and 28 days of treatment (that is, after 4 and 8 sessions of ECT; when the mean HRS-D global score was, respectively, 20.4 t 3.8 SD and 12.6 ? 2.5 SD). In the 15 patients who responded to treatment, a further determination was made immediately after complete clinical recovery (HRS-D global score of 4 or less, mean 2.1 + 0.7 SD), which occurred after 31-56 days of therapy (mean 42.5 & 8.6 SD) and required some additional ECT sessions, up to a maximum of 12. Throughout the study, patients did not receive any psychotropic drug (except low-dose benzodiazepines). Blood samples (30 ml) were drawn between 8:00 AM and 12:00 noon and were collected into plastic tubes containing 4 ml of anticoagulant (93 mu trisodium citrate, 213 mu citric acid, 111 mu glucose). Blood was centrifuged at room temperature for 20 min at 200 x g to obtain the platelet-rich plasma (PRP). The PRP was centrifuged at 16,000 x g for 10 min at 4°C and was washed twice with 30 ml buffer I (5 mu Tris-HCl ,20 mu Na*EDTA , 150 mu NaCl, pH 7.5). Platelet membranes were then prepared by hypotonic lysis in 30 ml buffer II (5 mu TrisHCl, 5 mu Na2EDTA, pH 7.5), homogenization, and centrifugation at 39,000 X g for 10 min. The pellet was washed with 30 ml buffer III (50 mu Tris-HCl, pH 7.4) and was resuspended in 30 ml incubation buffer (50 mu TrisHCl, 3 mu KCl, 120 mu NaCl, pH 7.4) at a
Brief Reports
concentration of 0.5- 1.2 mg protein/ml. For the determination of 3H-imipramine binding, an aliquot (180 ~1) of membrane suspension was incubated with 3H-imipramine (Amersham, Arlington Heights, IL; specific activity 21 Ci/mmol) at concentrations from 0.5 to 10 nh4 at 0°C for 60 min. Afterwards, a sample of 100 pl was taken, rapidly diluted in 5 ml of ice-cold buffer, and immediately filtered through Whatman GF/ B glass-fiber filters. The filters were washed with 3 X 5 ml of ice-cold buffer and dried, and the radioactivity was measured by liquid scintillation spectrometry. The specific binding of 3H-imipramine was defined as the difference between the total binding and that remaining in the presence of 100 pM desipramine. At 5 nrvf3H-imipramine, specific binding was about 70% of the total binding. The &, and the affinity constant (&) of 3Himipramine binding were calculated by linear regression of Scatchard plots. The B,, was expressed as fmohmg of platelet protein, as determined by the method of Lowry et aI. (195 1). Statistical analysis of data was made by Student’s t-test.
Results As shown in Table 1, mean baseline B, values of depressed patients were significantly lower than those of normal controls, whereas no significant difference between the two groups was observed with respect to mean Kd values. After 4 and 8 sessions of ECT, the mean density of platelet 3H-imipramine binding sites was slightly increased, but remained significantly lower than that of controls. At the time of complete clinical recovery, in patients who responded to treatment, mean B, values increased to the normal range. No effect of ECT on mean Kd values was observed.
Discussion The significant decrease of the density of platelet 3H-imipramine binding sites previously reported in depressed patients as compared with normal controls was confirmed by the present investi-
BIOL PSYCHIATRY 1988;24.469-472
Brief Reports
Table 1. Platelet 3H-ImipramineBinding in Depressed Patients before and after ECT and in Normal Controls 3H-Imipramine biding Subjects Depressed patients (n = 19) Before. ECT After 4 sessions of ECT After 8 sessions of ECT After complete clinical recovery+ Normal controls (n = 19)
B,
(fmoYmg protein)
& (W
486 f 12Sb
2.1 f 1.0
509 ? 15ob
2.0 f 0.9
550 f 157’
1.9 f 1.0
602 + 150
1.9 f 1.0
471
weeks of treatment with imipramine, our data demonstrate that the more rapid clinical effect of ECT as compared with tricyclic antidepressants is paralleled by a more rapid normalization of the density of platelet 3H-imipramine binding sites. This finding suggests a close link between the normalization of B, values and the biological events involved in the recovery from a depressive episode and supports the usefulness of platelet 3H-imipramine binding as a statedependent biological marker in depression.
References 647 f 134
Resultsare expressedas means
f
2.2 + 1.1
SD.
OData concerning responders to treatment (n =
15).
comparedwith the controlgroup. ‘p < 0.05 when companxlwith the control group.
“p < 0.01 when
gation. The mechanism responsible for this decrease remains unknown. Several authors, however, have recently suggested the existence of an endogenous inhibitor of the ‘H-imipramine binding site, which could act as a modulator of serotonin uptake, and whose concentration would change during a depressive episode, producing the decrease of B, values (Barbaccia et al. 1983; Langer et al. 1984; Rehavi et al. 1985). In our depressed patients, partial clinical improvement after 4 and 8 sessions of ECT was not followed by a normalization of B, values, a finding that is consistent with that reported by Langer et al. (1986) and is in line with the lack of a significant increase of the density of platelet 3H-imipramine binding sites observed by Suranyi-Cadotte et al. (1985) after partial clinical improvement following 2 weeks of treatment with irnipramine . On the other hand, the complete remission of depressive symptomatology (occurring in our responsive patients after 31-56 days of treatment) was followed by an increase of B, values to the normal range. As in the study of Suranyi-Cadotte et al. (1985), the normalization of B, values was manifested only after 8-12
Barbaccia ML, Gandolfi 0, Chuang DM, Costa E (1983): Modulationof neuronal serotonin uptake by a putative endogenous ligand of imipramine binding sites. Proc Nat1 Acad Sci USA 80:51345138. Briley M, Langer SZ, RaismanR, Sechter D, Zarifian E (1980): Tritiated imipramine binding sites are decreased in platelets of untreated depressed patients. Science 209:303-305. Fawcett J, Scheftner W (1986): Efficacy in depression: ECT versus antidepressants. Psychopharmacol Bull 22:468469.
Hamilton M (1960): A rating scale for depression. J Neurol Neurosurg Psychiatry 2356-62.
Kanof PD, Coccaro EF, Johns CA, Siever LJ, Davis KL (1987): Platelet (3H) imipramine binding in psychiatricdisorders.Biol Psychiatry 22:278-286. Langer SZ, RaismanR, Tahraoui L, Scatton B, Niddam R, Lee CR, Claustre Y (1984): Substituted tetrahydro-beta-carbolinesare possible candidates as endogenous ligand of the (3H) imipramine recognition site. Eur J Pharmacol98:153-154. Langer SZ, Sechter D, Loo H, Raisman R, Zarifian
E (1986): Electroconvulsive shock therapy and maximum binding of platelet titiated imipramine binding in depression. Arch Gen Psychiatry 43:949952. Lowry OH, Rosenbrough NJ, Farr AL, Randall RJ ( 1951): Protein measurements with the Folin phenol reagent. J Biol Chem 193:265-275.
Mellerup ET, Plenge P, Rosenberg R (1982): 3Himipramine binding sites in platelets from psychiatric patients. Psychiatry Res 7:221-227.
RaismanR, Sechter D, Briley MS, Zarifian E, Langer SZ (1981): High affinity 3H-imipramine binding in platelets from untreated and treated dearessed
472
BIOL PSYCHIATRY 1988;24:469-472
Brief Reports
patients compared to healthy volunteers. Psycho-
and serotonin uptake in depressed patients. Life
pharm4icology 75:368-371.
Sci 36:795-799.
Rehavi M, Ventura I, Same Y (1985): Demonstration of endogenous “imipramine-like” material in rat brain. Life Sci 36:686-693. Suranyi-Cadotte BE, Quiron R, Nair NPV, Lafaille F, Schwartz G (1985): Imipramine treatment differentially affects platelet 3H-imipramine binding
Wagner A, Aberg-Wistedt A, Asberg M, Eqvist B, Martensson B, Montero D (1985): Lower 3H-imipramine binding in platelets from untreated depressed patients compared to healthy controls. Psychiatry Res 16:131-139.
469
Changes in Platelet 3H-Imipramine Binding in Depressed Patients Receiving Electroconvulsive Therapy Mario Maj, Pasquale Mastronardi, Antonia Cerreta, Michele Romano, Basilio Mazzarella, and Dargut Kemali
Introduction A significant decrease in the density of platelet 3H-imipramine binding sites (B,) in untreated depressed patients as compared with healthy controls has been reported by several authors (Briley et al. 1980; Raisman et al. 1981; Wagner et al. 1985), although not confirmed by some others (Mellerup et al. 1982; Kanof et al. 1987). B, values have been found to remain low during partial clinical improvement following treatment with tricyclic antidepressants (Briley et al. 1980; Suranyi-Cadotte et al. 1985). but to increase to a normal range after 8-12 weeks of treatment, when complete symptomatic recovery is achieved (Suranyi-Cadotte et al. 1985). Similar results have been obtained by Langer et al. (1986) in depressed patients receiving electroconvulsive therapy (ECT): B,, values were still significantly lower than those of normal controls after 610 sessions of ECT, when clinical improvement was only partial, whereas they were found to be in the normal range after 12-l 8 months, at a time when patients were
From the Depltmcnt of Medical Psychology and Psycltiatty. Fmt Medical School. Uttivcnity of Nqks. Italy (MM., A.C., D.K.); theSewnd~of Araabcsidogy, Sccottd Medial S&ml. Uttivcnity of Nqh. Italy (P.M.. B.M.); atuI the Cam di Cm Via Chiant@. Nocen. hktno, Italy (MR.). AdmtsstqintmqtatstoPmfmuxhihohhj,Catt&adiIgiette Menule, Prima pOliclinica Univaritatio. Piazza Miraglia 2, I80138 Nqoli, Italy. Received Jttnc 30, 1987; revised !kphtbu 16. 1987.
0 1988 Society of Biological Psychiatry
euthymic. In this study, however, the 6 patients in whom platelet 3H-imipramine binding was determined after recovery from depression were on treatment with antidepressant drugs (4 of them also with lithium) at the time of this assessment, so that a possible effect of this additional treatment on B, cannot be excluded. Moreover, as the assessment was made several months after clinical improvement, it was impossible to explore whether or not the more rapid clinical effect of ECT in comparison with tricyclic antidepressants (Fawcett and Scheftner 1986) is paralleled by a more rapid normalization of platelet ‘H-imipramine binding. This last issue was addressed in the present investigation in which platelet 3H-imipramine binding was studied in a sample of patients with major depression at baseline, after 4 and 8 sessions of ECT, and immediately after recovery from the depressive episode.
Methods Nineteen inpatients (14 men and 5 women, age range 18-59 years, mean + SD 32.8 + 10.8), fulfilling DSM-III criteria for a diagnosis of major depressive episode, entered the study. They scored at least 16 (mean 25.7 & 4.2 SD) on the first 17 items of the 2 l-item version of the Hamilton Rating Scale for Depression @IRS-D) (Hamilton 1960) and had been free from psychotropic drugs (except low-dose benzodiaze-
470
BIOL PSYCHIATRY i9aa;24:469-472
pines) for at least 12 days. The control population included 19 subjects without personal or family history of psychiatric disorders (14 men and 5 women, age range 20-59 years, mean t SD 34.1 ? 12.0), who had not received psychotropic drugs in the previous month. Both patients and controls were physically healthy. Patients were treated with ECT because of the severity of depression or the unfavorable response to tricyclic antidepressants in previous episodes. Bilateral ECT was performed after premeditation with atropine sulfate, 1 mg im, thiopental sodium, 4 mg/kg iv, and succinylcholine chloride 0.7 mg/kg iv. Platelet 3H-imipramine binding was determined in patients and controls at baseline and in patients after 14 and 28 days of treatment (that is, after 4 and 8 sessions of ECT; when the mean HRS-D global score was, respectively, 20.4 t 3.8 SD and 12.6 ? 2.5 SD). In the 15 patients who responded to treatment, a further determination was made immediately after complete clinical recovery (HRS-D global score of 4 or less, mean 2.1 + 0.7 SD), which occurred after 31-56 days of therapy (mean 42.5 & 8.6 SD) and required some additional ECT sessions, up to a maximum of 12. Throughout the study, patients did not receive any psychotropic drug (except low-dose benzodiazepines). Blood samples (30 ml) were drawn between 8:00 AM and 12:00 noon and were collected into plastic tubes containing 4 ml of anticoagulant (93 mu trisodium citrate, 213 mu citric acid, 111 mu glucose). Blood was centrifuged at room temperature for 20 min at 200 x g to obtain the platelet-rich plasma (PRP). The PRP was centrifuged at 16,000 x g for 10 min at 4°C and was washed twice with 30 ml buffer I (5 mu Tris-HCl ,20 mu Na*EDTA , 150 mu NaCl, pH 7.5). Platelet membranes were then prepared by hypotonic lysis in 30 ml buffer II (5 mu TrisHCl, 5 mu Na2EDTA, pH 7.5), homogenization, and centrifugation at 39,000 X g for 10 min. The pellet was washed with 30 ml buffer III (50 mu Tris-HCl, pH 7.4) and was resuspended in 30 ml incubation buffer (50 mu TrisHCl, 3 mu KCl, 120 mu NaCl, pH 7.4) at a
Brief Reports
concentration of 0.5- 1.2 mg protein/ml. For the determination of 3H-imipramine binding, an aliquot (180 ~1) of membrane suspension was incubated with 3H-imipramine (Amersham, Arlington Heights, IL; specific activity 21 Ci/mmol) at concentrations from 0.5 to 10 nh4 at 0°C for 60 min. Afterwards, a sample of 100 pl was taken, rapidly diluted in 5 ml of ice-cold buffer, and immediately filtered through Whatman GF/ B glass-fiber filters. The filters were washed with 3 X 5 ml of ice-cold buffer and dried, and the radioactivity was measured by liquid scintillation spectrometry. The specific binding of 3H-imipramine was defined as the difference between the total binding and that remaining in the presence of 100 pM desipramine. At 5 nrvf3H-imipramine, specific binding was about 70% of the total binding. The &, and the affinity constant (&) of 3Himipramine binding were calculated by linear regression of Scatchard plots. The B,, was expressed as fmohmg of platelet protein, as determined by the method of Lowry et aI. (195 1). Statistical analysis of data was made by Student’s t-test.
Results As shown in Table 1, mean baseline B, values of depressed patients were significantly lower than those of normal controls, whereas no significant difference between the two groups was observed with respect to mean Kd values. After 4 and 8 sessions of ECT, the mean density of platelet 3H-imipramine binding sites was slightly increased, but remained significantly lower than that of controls. At the time of complete clinical recovery, in patients who responded to treatment, mean B, values increased to the normal range. No effect of ECT on mean Kd values was observed.
Discussion The significant decrease of the density of platelet 3H-imipramine binding sites previously reported in depressed patients as compared with normal controls was confirmed by the present investi-
BIOL PSYCHIATRY 1988;24.469-472
Brief Reports
Table 1. Platelet 3H-ImipramineBinding in Depressed Patients before and after ECT and in Normal Controls 3H-Imipramine biding Subjects Depressed patients (n = 19) Before. ECT After 4 sessions of ECT After 8 sessions of ECT After complete clinical recovery+ Normal controls (n = 19)
B,
(fmoYmg protein)
& (W
486 f 12Sb
2.1 f 1.0
509 ? 15ob
2.0 f 0.9
550 f 157’
1.9 f 1.0
602 + 150
1.9 f 1.0
471
weeks of treatment with imipramine, our data demonstrate that the more rapid clinical effect of ECT as compared with tricyclic antidepressants is paralleled by a more rapid normalization of the density of platelet 3H-imipramine binding sites. This finding suggests a close link between the normalization of B, values and the biological events involved in the recovery from a depressive episode and supports the usefulness of platelet 3H-imipramine binding as a statedependent biological marker in depression.
References 647 f 134
Resultsare expressedas means
f
2.2 + 1.1
SD.
OData concerning responders to treatment (n =
15).
comparedwith the controlgroup. ‘p < 0.05 when companxlwith the control group.
“p < 0.01 when
gation. The mechanism responsible for this decrease remains unknown. Several authors, however, have recently suggested the existence of an endogenous inhibitor of the ‘H-imipramine binding site, which could act as a modulator of serotonin uptake, and whose concentration would change during a depressive episode, producing the decrease of B, values (Barbaccia et al. 1983; Langer et al. 1984; Rehavi et al. 1985). In our depressed patients, partial clinical improvement after 4 and 8 sessions of ECT was not followed by a normalization of B, values, a finding that is consistent with that reported by Langer et al. (1986) and is in line with the lack of a significant increase of the density of platelet 3H-imipramine binding sites observed by Suranyi-Cadotte et al. (1985) after partial clinical improvement following 2 weeks of treatment with irnipramine . On the other hand, the complete remission of depressive symptomatology (occurring in our responsive patients after 31-56 days of treatment) was followed by an increase of B, values to the normal range. As in the study of Suranyi-Cadotte et al. (1985), the normalization of B, values was manifested only after 8-12
Barbaccia ML, Gandolfi 0, Chuang DM, Costa E (1983): Modulationof neuronal serotonin uptake by a putative endogenous ligand of imipramine binding sites. Proc Nat1 Acad Sci USA 80:51345138. Briley M, Langer SZ, RaismanR, Sechter D, Zarifian E (1980): Tritiated imipramine binding sites are decreased in platelets of untreated depressed patients. Science 209:303-305. Fawcett J, Scheftner W (1986): Efficacy in depression: ECT versus antidepressants. Psychopharmacol Bull 22:468469.
Hamilton M (1960): A rating scale for depression. J Neurol Neurosurg Psychiatry 2356-62.
Kanof PD, Coccaro EF, Johns CA, Siever LJ, Davis KL (1987): Platelet (3H) imipramine binding in psychiatricdisorders.Biol Psychiatry 22:278-286. Langer SZ, RaismanR, Tahraoui L, Scatton B, Niddam R, Lee CR, Claustre Y (1984): Substituted tetrahydro-beta-carbolinesare possible candidates as endogenous ligand of the (3H) imipramine recognition site. Eur J Pharmacol98:153-154. Langer SZ, Sechter D, Loo H, Raisman R, Zarifian
E (1986): Electroconvulsive shock therapy and maximum binding of platelet titiated imipramine binding in depression. Arch Gen Psychiatry 43:949952. Lowry OH, Rosenbrough NJ, Farr AL, Randall RJ ( 1951): Protein measurements with the Folin phenol reagent. J Biol Chem 193:265-275.
Mellerup ET, Plenge P, Rosenberg R (1982): 3Himipramine binding sites in platelets from psychiatric patients. Psychiatry Res 7:221-227.
RaismanR, Sechter D, Briley MS, Zarifian E, Langer SZ (1981): High affinity 3H-imipramine binding in platelets from untreated and treated dearessed
472
BIOL PSYCHIATRY 1988;24:469-472
Brief Reports
patients compared to healthy volunteers. Psycho-
and serotonin uptake in depressed patients. Life
pharm4icology 75:368-371.
Sci 36:795-799.
Rehavi M, Ventura I, Same Y (1985): Demonstration of endogenous “imipramine-like” material in rat brain. Life Sci 36:686-693. Suranyi-Cadotte BE, Quiron R, Nair NPV, Lafaille F, Schwartz G (1985): Imipramine treatment differentially affects platelet 3H-imipramine binding
Wagner A, Aberg-Wistedt A, Asberg M, Eqvist B, Martensson B, Montero D (1985): Lower 3H-imipramine binding in platelets from untreated depressed patients compared to healthy controls. Psychiatry Res 16:131-139.