Initial serotonin transport into viable platelets and imipramine binding to platelet membranes

Summary. Transport of serotonin into human platelets is a paradigm for neuronal reuptake to investigate putatively low neurotransmitter availability in certain psychiatric diseases. However, inconsistent results have been obtained on serotonin binding to platelet membranes at equilibrium and transport during initial phase into isolated platelets. In the present study we applied a rapid oil-centrifugation technique to study ¹⁴C-serotonin transport for 15 s into viable human platelets during the initial phase, compared to the binding of ³H-imipramine at equilibrium (60 min) to membranes isolated from platelets of the same individuals and to their blood serotonin levels. Platelets were viable for two h after the isolation procedure; concomitant with the decrease in viability transport also decreased. Initial transport into viable cells was observed for two min. Across 19 healthy individuals blood serotonin levels correlated with the halfmaximal saturation constants of binding, K_D, for imipramine but not with any other transport or binding parameters, such as V_max or B_max. Inhibition studies with psychoactive drugs showed good correlation between transport during the initial phase and binding at equilibrium (r = 0.83). It is speculated that changes in the V_max of transport reflect problems with isolation, pretreatment with drugs, the energy load of the cell, and polymorphism of the serotonin transporter. The latter shows a polymorphism in the 5′regulatory region with a 44-bp insertion (l, long form) or deletion (s, short form). Results by Greenberg et al. indicate that in platelets from healthy men the l-variant was associated with increased initial serotonin uptake. Thus for genotyping, we suggest to subdivide patient and control groups in addition to psychopathology also according to their peripheral biochemical lesions. Received December 3, 2001; accepted February 4, 2002     

High-affinity imipramine binding and serotonin uptake in platelets of eight adolescent and ten adult obsessive-compulsive patients

The authors evaluated high-affinity [3H]imipramine binding and [3H]serotonin uptake to platelets in eight adolescent and 10 adult patients who met DSM-III criteria for obsessive-compulsive disorder in comparison with those of normal control subjects of similar ages. The maximal binding of [3H]imipramine was significantly lower in adults and adolescents with obsessive-compulsive disorder than in the control subjects. No differences between groups in the affinity of [3H]imipramine to its binding sites or in serotonin uptake kinetic measures were detected. The lower density of [3H]imipramine binding sites in platelet membrane in patients with obsessive-compulsive disorder might implicate involvement of the serotonergic system or might represent an adaptive response to a chronic disease.     

Reduced platelet serotonin in depression

Platelet serotonin levels were measured in several psychiatric disorders to determine whether they distinguish among major depressive disorder (one or more depressive episodes and no manic episodes), dysthymic disorder (depressive neurosis), and schizophrenic and paranoid disorders. Serotonin levels in 141 subjects were determined using high performance liquid chromatography with electrochemical detection. Serotonin (5HT) levels in control subjects were significantly lower in males than in females. A marked reduction in 5HT levels, as compared to controls, was found in male and female patients with major depressive disorder, but not in dysthymic disorder. A slight but significant reduction in serotonin levels was found in female schizophrenic patients. The reduction in serotonin levels found in major depressive disorder could not be attributed to chronic antidepressant treatment. Liquid chromatography with electrochemical detection used in the present study permits a large-scale investigation.     

Effects of chlorpromazine on serotonin uptake in blood platelets

Platelet serotonin (5-HT) uptake was studied in schizophrenic patients before and after treatment with chlorpromazine (CPZ) for 2-3 weeks. No difference was noted in the affinity of 5-HT (Km) or maximum velocity of 5-HT uptake (Vmax) of unmedicated schizophrenic patients and normal controls. Administration of CPZ was associated with a significant increase in Km and Vmax. Specific uptake of 5-HT, at 0.5 microM, was significantly decreased in most subjects. Chlorpromazine inhibited 5-HT uptake into platelets from normal controls in vitro (IC50, 1.8 +/- 0.1 microM) in a competitive manner.     

Platelet imipramine binding and serotonin uptake in obsessive-compulsive patients

Platelet imipramine binding was measured in 16 drug-free nondepressed patients (aged 20-61 years, mean ± SD 35 ± 8) suffering from obsessive-compulsive disorder (OCD) and in 16 sex-, race- and age-matched healthy controls. Imipramine binding capacity and affinity were not different in the 2 groups. Platelet serotonin (5-HT) uptake capacity, V_max, was also measured in 15 of these patients and their matched controls. V_max was significantly higher in the patients (309 ± 149 pmol/10⁹ cells/min) than in the controls (181 ± 110). An increase in platelet 5-HT uptake supports the involvement of 5-HT in OCD and may suggest that a hyperactive serotonergic system is present in this disorder.     

Reduced whole blood serotonin in major depression

Whole blood serotonin (WBS) was measured in 17 patients with DSM-III-R major depression and compared to a healthy control group of 57. Values were significantly lower in the depressed group, but there was no correlation with the degree of depression. Four patients with a history of suicide attempts had even lower levels, but this was not statistically significant. Across all subjects, there was an effect of season of sampling, with values in spring significantly higher than those in autumn. Since blood serotonin is primarily produced peripherally, these results suggest that some aspect of peripheral serotonin metabolism is abnormal in major depression. Depression and Anxiety 5:108–111, 1997. © 1997 Wiley-Liss, Inc.     

Influence of age, sex and diurnal variability on imipramine receptor binding and serotonin uptake in platelets of normal subjects.

Imipramine (IMI) binding and serotonin (5-HT) uptake were determined in platelets of 98 healthy volunteers; and their association with age, sex and circadian rhythm were evaluated. A large interindividual variability was found for both IMI and 5-HT parameters. There was a negative correlation of IMI affinity constant (Kd) and binding (Bmax) with age, but no such correlation of 5-HT affinity constant (Km) or uptake (Vmax). Significant age-related diurnal variability was found for 5-HT Km in the whole group as well as for IMI Kd in males, but not in females. There was no significant correlation between 5-HT Vmax and IMI Bmax. Our results underscore a cautious approach to the interpretation of platelet serotonergic studies. In light of the multiple variables influencing the results, the usefulness of IMI or 5-HT as clinical markers should be re-evaluated.     

Effects of lithium carbonate on serotonin uptake in rat blood platelets

1. Lithium carbonate administration to male rats for 1 to 4 weeks increased the affinity of platelet serotonin (5-HT) uptake (decreased Km) and decreased the rate of 5-HT uptake (Vmax) when the serum lithium levels were 0.5 meq/1 to 0.65 meq/l. When the serum levels were increased further to 1 meq/1, Vmax decreased further, without additional change in Km. 2. No effect on the kinetics of 5-HT uptake was observed when serum lithium levels were at 0.4 meq/1 or below. 3. Addition of lithium carbonate in vitro (1 meq/1) had no effect on the Km and Vmax of rat platelet 5-HT uptake. 4. The possible mechanism of the inhibitory effect of lithium treatment on platelet 5-HT uptake is discussed.     

Marked reduction in the number of platelet-tritiated imipramine binding sites in geriatric depression

The number (Bmax) and affinity (Kd) of platelet-tritiated imipramine binding sites was determined in young and middle-aged controls 50 years of age and younger (n = 25), elderly normal controls over 60 years of age (n = 18), patients who fulfilled DSM-III criteria for major depression who were under 50 years of age (n = 29), patients who fulfilled DSM-III criteria for major depression who were 60 years of age and older (n = 19), and patients who fulfilled both DSM-III criteria for primary degenerative dementia and National Institute of Neurological and Communicative Disorders and Stroke-Alzheimer's Disease and Related Disorders Association criteria for probable Alzheimer's disease (n = 13). Both groups of depressed patients (under 50 and over 60 years of age) exhibited significant reductions (decreases 42%) in the number of platelet-tritiated imipramine binding sites with no change in affinity, when compared with their age-matched controls. There was little overlap in Bmax values between the elderly depressed patients and their controls. The patients with probable Alzheimer's disease showed no alteration in platelet-tritiated imipramine binding. There was no statistically significant relationship between postdexamethasone plasma cortisol concentrations and tritiated imipramine binding. These results indicate that platelet-tritiated imipramine binding may have potential utility as a diagnostic adjunct in geriatric depression, and moreover that the reduction in the number of platelet-tritiated imipramine binding sites is not due to hypercortisolemia.     

Reduced serotonin content and uptake in platelets from patients with essential hypertension: is a ouabain-like factor involved?

Platelet serotonin (5-HT) content was investigated in 35 essential hypertensive patients and in 58 normotensive subjects. A significant decrease of platelet 5-HT content was observed in both hypertensive men and women in respect to normotensive controls. This decreased platelet 5-HT content appears to be linked to a reduced capacity of platelets to take up 5-HT. Maximal velocity of 5-HT uptake was shown to be reduced in platelets from hypertensive patients and significantly correlated to platelet 5-HT content. No concomitant change of uptake Michaelis constant (KM) was observed. The great sensitivity of human platelets to very low concentrations of ouabain was demonstrated when incubation times were prolonged. The decreased Vmax observed in platelets from hypertensive patients and reproduced by ouabain inhibition could conceivably be linked to the presence of a circulating ouabain-like factor in hypertension.     

Upregulation of imipramine binding and serotonin uptake by estradiol in female rat brain

This report describes the effect of chronic estradiol treatment on the serotonin transporter in the female rat brain. Both [3H]imipramine binding and [3H]serotonin uptake increased by 20-30% in the frontal cortex and hypothalamus of ovariectomized rats after 12 days of 17 beta-estradiol treatment. No differences were observed in the binding and uptake parameters as a function of the rats' estrous cycle or in untreated ovariectomized rats, as compared to controls. Estradiol in vitro, inhibits [3H]imipramine binding as well as serotonin uptake in rat brain and human platelets. Like serotonin, estradiol decreases the dissociation rate in vitro of [3H]imipramine from its binding site in a dose-dependent manner.     

The stereoselectivity of serotonin uptake in brain tissue and blood platelets: the topography of the serotonin uptake area

This review concerns effects of stereoisomers on 5-HT uptake in brain tissue and/or blood platelets. All studies in which at least a pair of stereoisomers were used are considered. Differences between effects of stereoisomers of antidepressants as well as other drugs on 5-HT uptake are discussed. The findings indicate that 5-HT uptake is a stereoselective process. A topographical model of the 5-HT uptake area is proposed, based mainly on comparisons between spatial features of stereoisomers that inhibit 5-HT uptake.     

Serotonin uptake and imipramine binding in the blood platelets of obsessive-compulsive disorder patients.

14C-Serotonin (5-HT) uptake and 3H-imipramine binding (IB) were studied in the blood platelets of 20 obsessive-compulsive disorder (OCD) patients, 53 normal controls (5-HT uptake) and 32 normal controls (IB binding). The maximum number of binding sites (Bmax) was significantly decreased in OCD patients compared to normal controls, but there was no difference in the affinity for 3H-imipramine (Kd). The affinity for 5-HT uptake (Km) was also decreased in the OCD patients but the maximum velocity of 5-HT uptake sites (Vmax) was not significantly different in OCD patients and normal volunteers. There were trends for the Slowness Subscale of the Maudsley Obsessional-Compulsive Inventory (MOCI) to be positively correlated with the Km of 5-HT uptake (p = 0.094), whereas the Global Scale, Checking Subscale, and Doubting Conscientiousness Subscale of MOCI were negatively correlated with the Kd of IB (p = 0.066, p = 0.08, and p = 0.062, respectively). The results provide further evidence for the dysfunction of the serotonergic system in OCD.     

Allelic differences in the serotonin transporter-linked polymorphic region in geriatric depression.

Previous studies have examined the role of genetic variations in the serotonin transporter-linked polymorphic region (5HTTLPR) in affective disorders. The authors studied 182 older depressed subjects and 107 elderly control subjects and obtained DNA for genotyping at the 5HTTLPR. There were no significant differences in allele frequencies generally or for number of short alleles for the group as a whole, but interesting gender effects emerged. Among men, 23% of depressed men had two short alleles, compared with only 5% of control subjects. Among women, 67% of depressed women with more than one episode had at least one short allele, compared with 41% of single-episode female patients. Also, 74% of women with a positive family history of psychiatric illness in any female relative had at least one short allele, whereas 53% had at least one short allele who did not have such a family history. Our results add to the literature linking this gene to affective illness. The negative association of allele frequency and depression may be related to the relatively small sample size. The findings raise the possibility that this genetic locus may exert differential effects based on gender, increasing risk in men, and increasing risk of recurrence in women.     

Aberrant seasonal variations of platelet serotonin uptake in endogenous depression

The serotonin uptake in platelets of 120 healthy volunteers and 64 endogenously depressed patients was investigated over a 2-year period. In healthy individuals, Km exhibited a significant seasonal rhythm during the bright half of the year. The seasonal rhythm of Vmax assumes the form of a sine curve, with nadir values at the vernal and autumn equinoxes and peak values at the winter and summer solstices. Km in patients was higher than in controls in February and October, and the seasonal variation of Km differed between patients and controls. The monthly mean values of Vmax in patients were, as a rule, lower than corresponding values in controls, but significantly so only in December. Patients had higher Vmax than controls in October and November, and the seasonal variation of Vmax in patients differed from that of controls. The results suggest that Km, a measure of the affinity of the serotonin uptake site, may be subject to photoperiodic regulation in healthy individuals. The annual variation in uptake site densities, as judged by the changes in Vmax, are probably generated by an endogenous superior oscillator. The aberrant uptake kinetics found in the endogenously depressed patients may reflect seasonal susceptibility to the disorder and/or altered serotonergic rhythmicity.     

Genetic control of serotonin uptake in blood platelets: a twin study

Platelet serotonin (5HT) uptake was studied in 13 pairs of monozygotic (MZ) twins, 13 pairs of dizygotic (DZ) twins, and 14 pairs of unrelated normal volunteers. Significant intraclass correlations (ICC) in the affinity (Km) of 5HT uptake in the blood platelets of MZ and DZ twins and unrelated pairs were found. However, the ICC for maximum velocity (Vmax) was significant only in MZ and DZ twins. The ICC of the Vmax of 5HT uptake of MZ twins was significantly greater than that of DZ twins and unrelated pairs. This suggests that the Vmax of 5HT uptake in blood platelets is, in part, heritable. Thus, low platelet 5HT uptake (Vmax) in major depression and other disorders may be genetically determined.