Per Plenge

High affinity binding of [3H]paroxetine and [3H]imipramine to human platelet membranes.

Paroxetine, one of the most potent and specific serotonin uptake inhibitors, was tritiated and used for binding studies with human platelet membranes. Specific, high affinity binding was demonstrated. The binding was compared with [3H]imipramine binding; it was found that the maximal binding (Bmax) was the same for [3H]paroxetine and [3H]imipramine, whereas the affinity was much higher for [3H]paroxetine (KD 0.08 nM and 0.56 nM for paroxetine and imipramine binding, respectively). IC50 was calculated for the inhibition of [3H]paroxetine and [3H]imipramine binding by a number of antidepressants; the corresponding Hill coefficients were also calculated.

3H-imipramine binding sites in platelets from psychiatric patients

Abstract High-affinity binding of 3 H-imipramine to platelet membranes from control subjects and psychiatric patients was studied. Maximal binding (βmax) in depressed manic-melancholic patients was higher than in euthymic manic- melancholic patients, psychiatric controls, and normal controls.

Brain 5-HT1A, 5-HT1D, and 5-HT2 Receptors in suicide victims

We report on 5-HT1A, 5-HT1D, and 5-HT2 binding sites in 23 control subjects and 18 suicide victims subdivided according to the method of death and the previous existence of depressive symptoms. No difference in maximum binding (Bmax) or binding affinity (Kd) was found between the control and overall suicide groups for the binding sites studied. The drug overdose subgroup showed, however, a significant decrease in the 5-HT1A binding affinity, probably explained by the higher sensitivity of this binding site to the acute administration of tricyclic antidepressants. A significant decrease in 5-HT1D binding affinity was also found in the depressed suicides, together with a significant decrease in the number of 5-HT1D binding sites in the nondepressed suicides. Further studies should be carried out on the 5-HT1D binding site as it might represent a new tool in the understanding of the depressive illness.

High affinity binding of 3H-paroxetine and 3H-imipramine to rat neuronal membranes.

Paroxetine is the most potent and one of the most specific serotonin uptake inhibitors. High-affinity3H-paroxetine and3H-imipramine binding was compared in rat neuronal membranes.TheKd value for3H-paroxetine binding to neuronal membranes was 0.08 nM, which is exactly the same value as with platelet membranes. TheKd value for3H-imipramine binding to neuronal membranes was about 4 nM, which is higher than theKd value for3H-imipramine binding to platelet membranes (0.5 nM).The results indicated that the3H-paroxetine binding site is identical in neuronal membranes and in platelet membranes; this binding site is probably located on the serotonin transport mechanism. In addition, part of the highaffinity3H-imipramine binding to neuronal membranes is probably located on the serotonin transport mechanism, but another part is located elsewhere. Furthermore the polypeptides containing the3H-imipramine binding sites may not be identical in neuronal and platelet membranes.

Characterization of an allosteric citalopram-binding site at the serotonin transporter

The serotonin transporter (SERT), which belongs to a family of sodium/chloride‐dependent transporters, is the major pharmacological target in the treatment of several clinical disorders, including depression and anxiety. In the present study we show that the dissociation rate, of [3H]S‐citalopram from human SERT, is retarded by the presence of serotonin, as well as by several antidepressants, when present in the dissociation buffer. Dissociation of [3H]S‐citalopram from SERT is most potently inhibited by S‐citalopram followed by R‐citalopram, sertraline, serotonin and paroxetine. EC50 values for S‐ and R‐citalopram are 3.6 ± 0.4 µm and 19.4 ± 2.3 µm, respectively. Fluoxetine, venlafaxine and duloxetine have no significant effect on the dissociation of [3H]S‐citalopram. Allosteric modulation of dissociation is independent of temperature, or the presence of Na+ in the dissociation buffer. Dissociation of [3H]S‐citalopram from a complex with the SERT double‐mutant, N208Q/N217Q, which has been suggested to be unable to self‐assemble into oligomeric complexes, is retarded to an extent similar to that found with the wild‐type, raising the possibility that the allosteric mechanism is mediated within a single subunit. A species‐scanning mutagenesis study comparing human and bovine SERT revealed that Met180, Tyr495 and Ser513 are important residues in mediating the allosteric effect, as well as contributing to high‐affinity binding at the primary site.

Effect of aging in human cortical pre- and postsynaptic serotonin binding sites

5-HT1A, 5-HT1D, 5-HT2 binding sites and affinity and 5-HT uptake sites were simultaneously determined in frontal cortex samples from 23 control subjects, aged 16-75 years. A significant reduction in the number of 5-HT1D and 5-HT2 binding sites was found with regard to age, together with a significant decrease in the 5-HT2 binding affinity. It is suggested that the total 5-HT1 age-related loss described in previous studies could be ascribed to the 5-HT2 subtype. Furthermore, aging does not seem to be associated with a reduced cortical serotonergic innervation, as indicated by the stability of the [3H]paroxetine-labeled 5-HT uptake sites.

Antidepressive drugs can change the affinity of [3H]imipramine and [3H]paroxetine binding to platelet and neuronal membranes

Serotonin transport in synapses and platelets is inhibited by tricyclic antidepressants as well as by more selective transport inhibitors. This inhibition is hypothesized to be of importance for the psychotropic effect, although it is known that some new antidepressants do not possess this transport inhibitory action. We now report that antidepressive drugs can influence the serotonin transport complex in platelets and brain in other ways: [3H]imipramine and [3H]paroxetine, which bind with high affinity to the serotonin transport complex, can be dissociated from the complex with velocity constants strongly influenced by the different antidepressants. This effect is not correlated to the inhibitory action of the drugs on serotonin transport. Furthermore the effect is seen in the micromolar range in contrast to the high affinity binding process which takes place in the pico- and nanomolar range. The effects of antidepressants on the dissociation rates of bound ligand make it possible to differentiate between serotonin reuptake inhibitors which appear identical in other assays. Antidepressive drugs can thus be divided into groups which differ from the usual classifications.

Affinity modulation of [3H]imipramine, [3H]paroxetine and [3H]citalopram binding to the 5-HT transporter from brain and platelets.

The dissociations of [3H]imipramine, [3H]paroxetine and [3H]citalopram from the 5-HT (serotonin 5-hydroxytryptamine) transporter were found to be markedly influenced by several drugs, although concentrations in the microM range were needed. Most of these drugs attenuated the dissociation rate, i.e. increased the affinity between the ligand and the binding site. A few increased the dissociation rate however. The binding of drugs to the affinity-modulating site was specific, although of low affinity and probably changing the conformation of the high-affinity binding site, thereby changing the fit between the ligand and the interacting amino acid side-chains. Although the drugs usually affected the dissociation rates of the three ligands in the same manner, there were some which had different effects on [3H]imipramine, [3H]paroxetine and [3H]citalopram. For example, 5-HT markedly attenuated the dissociation of [3H]imipramine, had a moderate effect on [3H]paroxetine and very little effect on [3H]citalopram dissociation. This indicates that the three ligands are bound to different domains on the 5-HT transporter. [3H]Citalopram dissociation from human brain and rat brain were differently affected by several drugs. Indalpine augmented the dissociation rate of the [3H]citalopram 5-HT transport complex in human brain but attenuated it in rat brain, thus revealing a species difference of the 5-HT transporter.

Regional distribution of the serotonin transport complex in human brain, identified with 3H-Paroxetine, 3H-Citalopram and 3H-Imipramine

1. Regional distribution of the serotonin transport complex was studied in 12 different brain areas from human brains. The serotonin uptake complex was measured with 3H-paroxetine, and 3H-imipramine. The binding site density was highest in the nucleus of raphé, medium in the basal ganglia, and lowest in cortical areas. The specific binding measured with 3H-paroxetine and 3H-citalopram was compared with the high affinity 3H-imipramine binding determined with either 100 microM 5HT or 1 microM imipramine as non specific displacers. 3H-paroxetine and 3H-citalopram allowed a more precise determination of Bmax, and are both good ligands for the serotonin uptake site, but the determinations with 3H-imipramine were within the same range. 2. Protease digestion of brain membranes showed that the binding site measured with all three ligands disappeared with the same rate as other membrane proteins, and not faster as might be expected from the literature. 3. Left/right hemisphere distribution was measured in cortical tissue from 6 brains using 3H-paroxetine. No difference between the two hemispheres was found. In one brain from a lithium treated patient a very low binding was measured, possibly indicating that the lithium treatment had decreased the serotonin uptake mechanism.

Lithium treatment: does the kidney prefer one daily dose instead of two?

Renal structure and function were investigated in two groups of long‐term lithium treated patients. Lithium was administered in two different ways either in a one‐dose per day schedule where the whole dose of lithium was given between 8 and 10 p.m. or in a schedule where the lithium dose was given, divided into two or three doses, during the day. Kidney biopsy was performed, and structural changes in the kidney tissue were determined together with 24‐h urine volume in the individual patients. The functional as well as the structural changes were most pronounced in patients given their lithium in divided doses during the day. Lithium may be more harmful to the kidney when the lithium administration gives a relatively constant serum lithium level than when the administration causes greater variations including peak values and low minimum levels in serum lithium. The reason for this might be that a number of regenerative processes only occur in periods with low lithium concentrations.