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Objective: To investigate the in vitro and in vivo effects of 5-hydroxytryptamine (5-HT) on the rat urinary bladder and to characterize the receptors involved in mediating these pharmacological effects by using selective antagonists., Materials and Methods: Female Wistar rats (250-350 g) were used for all studies. In vitro, detrusor muscle strips were mounted between two platinum electrodes in organ baths filled with a modified Krebs' solution bubbled with 95% O(2) and 5% CO(2) at 37 degrees C. After equilibration and a contraction to 80 mmol/L KCl, strips were exposed to electrical field stimulation for 30 min and incubated with the antagonist or vehicle for a further 30 min, then a 5-HT concentration-response curve (CRC) was obtained. In vivo, rats were anaesthetized with pentobarbital, and the ureters and urethra ligated, the bladder catheterized and infused with saline. 5-HT (3-100 microg/kg intravenous) dose-dependently increased intravesical pressure (IVP). After administering 5-HT at 30 microg/kg three times at 10 min intervals (controls), one dose of antagonist was perfused for 5 min and, after a further 5 min, 30 microg/kg 5-HT was tested again. This cycle was repeated four times using increasing doses of the antagonist to be tested., Results: In vitro, 5-HT (0.01-100 micromol/L) induced a concentration-dependent enhancement of the neurogenic response, with a mean (sd) pEC(50) of 6.36 (0.15) and E(max) of 41.1 (4.6)% KCl (eight rats). In unstimulated tissues, 5-HT induced no contractile effect. Selective 5-HT(4), 5-HT(3) and 5-HT(1A) receptor antagonists had no effect on the 5-HT potentiating effects. The potentiating effect of 5-HT was antagonized by mesulergine at 0.3 micromol/L, R(+)lisuride at 0.3 micromol/L and the selective 5-HT(7) receptor antagonist SB-258741 at 0.3 micromol/L. In vivo, in anaesthetized rats, IVP increases induced by repeated doses of 30 microg/kg 5-HT were reproducible. R(+)lisuride (3-100 microg/kg) dose-dependently inhibited the 5-HT-induced increase of IVP. At the maximum dose tested, R(+)lisuride almost totally inhibited the 5-HT effect., Conclusions: In rat isolated detrusor muscle the 5-HT(7) receptor antagonists SB-258741, R(+)lisuride and mesulergine blocked the 5-HT potentiating effect with the expected potency. Moreover, in anaesthetized rats, R(+)lisuride abolished 5-HT effects on IVP at doses that antagonize physiological effects known to be mediated by 5-HT(7) receptor activation in several animal species. These results suggest the involvement of 5-HT(7) receptors in the modulation of rat bladder contraction both in vitro and in vivo.

Quinolin-2-ones bearing a heteroaryl-piperazine linked by a two carbon chain at the 3- or 4-position were synthesised and evaluated as mixed 5-HT(1B)/5-HT(2A) receptor antagonists. Potent mixed antagonists were obtained with thieno[3,2-c]pyridine derivatives. In this series, compound 2.1 (SL 65.0472) proved to be functional antagonist at both the 5-HT(2A) receptor (rat in vivo 5-HT-induced hypertension model) and the 5-HT(1B) receptor (dog in vitro saphenous vein assay).

In the rat brain, the presynaptic 5-hydroxytryptamine (5-HT) autoreceptors located on 5-HT terminals correspond to the 5-HT1B subtype. The presence of a 5-HT receptor probably located on 5-HT nerve endings and modulating transmitter release in the human neocortex has been reported, but its detailed pharmacological characterization is not yet available. On the other hand, receptor binding and autoradiographic results indicate that the 5-HT1B receptor subtype is not present in the human brain. We, therefore, studied the modulation of the electrically evoked release of [3H]5-HT by various 5-HT receptor agonists and antagonists in preloaded slices of human neocortex obtained from 18 patients undergoing neurosurgery. The nonselective 5-HT1A/1B/1D receptor agonist 5-carboxamidotryptamine produced a potent inhibition (70% at 0.03 microM) of the electrically evoked release of [3H]5-HT which was blocked by 5-HT receptor antagonists with the following relative order of potency: methiothepin greater than metergoline = methysergide greater than propranolol. The selective 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin at 0.1 microM did not modify the electrically evoked release of [3H]5-HT. The 5-HT1A/1B receptor agonist RU 24969 was 10 times more potent at inhibiting [3H]5-HT overflow in the rat frontal cortex than in the human neocortex. The potent 5-HT1B receptor antagonist cyanopinodolol did not modify the 5-carboxamidotryptamine-induced inhibition of the electrically evoked release of [3H]5-HT in slices of the human neocortex, but produced by itself a small inhibition of [3H]5-HT overflow.(ABSTRACT TRUNCATED AT 250 WORDS)

[3H]-Imipramine and [3H]-paroxetine label with high affinity a site which is associated with the serotonergic transporter in brain and platelets. The pharmacological profile of inhibition by drugs of [3H]-imipramine and [3H]-paroxetine binding is highly correlated with the potency of the drugs to inhibit the uptake of 5HT. Denervation of serotonergic neurons by electrolytic lesions or with 5,7-dihydroxytryptamine produces marked decreases in the density of [3H]-imipramine as well as [3H]-paroxetine binding. Dissociation kinetic experiments support the view that the substrate recognition site for 5HT is different from the modulatory site which is labelled by [3H]-imipramine or [3H]-paroxetine. The existence of an endogenous ligand acting on the [3H]-imipramine recognition site to modulate the 5HT transporter was proposed by several laboratories. [3H]-Imipramine binding in platelets appears to be a biological marker in depression. Studies carried out in several laboratories report a significant decrease in the Bmax of platelet [3H]-imipramine binding without changes in Kd, when severely depressed untreated patients are compared with healthy volunteers matched for age and sex. The Bmax of platelet [3H]-imipramine binding appears to be a state-dependent biological marker in depression. It is tempting to speculate that the endocoid of the [3H]-imipramine recognition site may play a role in the pathogenesis of depression.

The 5-hydroxytryptamine (5HT) receptor agonist, 5-methoxytryptamine, inhibited in a concentration-dependent manner the electrically-evoked release of 3H-5HT from superfused rat hypothalamic slices, with an IC50 of 560 nmol/l, without affecting the spontaneous outflow of radioactivity. In the presence of the selective monoamine oxidase B (MAO B) inhibitor, (-)-deprenyl (1 mumol/l), the concentration-effect curve for 5-methoxytryptamine was shifted significantly to the left, and the IC50 was decreased to 25 nmol/l. Under the same experimental conditions, the potency of the 5HT receptor agonist lysergic acid diethylamide (LSD) at inhibiting the electrically-evoked release of 3H-5HT was the same in the presence as well as in the absence of (-)-deprenyl. The IC50 values for LSD were 34 nmol/l in the absence of deprenyl, and 31 nmol/l in the presence of the MAO B inhibitor. It is concluded that deprenyl potentiates the inhibition by 5-methoxytryptamine of 3H-5HT release, by preventing its inactivation through MAO B. Since 5-methoxytryptamine may be present in the pineal gland of some species, the potent effects of this 5-HT receptor agonist on serotoninergic neurotransmission may be of physiological relevance.

The overflow of tritium from stimulated rat hypothalamic slices preincubated with [3H]-serotonin (5-HT) was significantly enhanced by reducing the frequency of stimulation from 3 Hz to 1 Hz while keeping the number of impulses constant. The 5-HT receptor agonist 5-methoxytryptamine inhibited in a concentration-dependent manner the electrically-evoked release of [3H]-5-HT with IC50 values of 560 nmol/l and of 34 nmol/l when the stimulations were delivered at 3 Hz and 1 Hz, respectively. The terminal 5-HT autoreceptor antagonist methiothepin enhanced in a concentration-dependent manner the electrically-evoked release of [3H]-5-HT and this effect was greater at a frequency of stimulation of 3 Hz than at 1 Hz. In the same paradigm, the 5-HT reuptake inhibitors citalopram and paroxetine did not alter the overflow of radioactivity elicited by stimulation at 3 Hz but significantly decreased it at 1 Hz. In the presence of 5-HT autoreceptor blockade achieved with methiothepin, citalopram increased the overflow of [3H]-5-HT to the same extent at 1 Hz and at 3 Hz. The IC50 values for inhibition of [3H]-5-HT release by the selective alpha 2-adrenoceptor agonist UK 14.304 were 35 nmol/l at 3 Hz and 30 nmol/l at 1 Hz. It is concluded that modulation of 5-HT release by 5-HT autoreceptors, but not by alpha 2-adrenoceptors is dependent on the synaptic concentration of 5-HT as a function of the frequency of depolarization.

In slices of the rat hypothalamus prelabeled with [3H]-5-hydroxytryptamine [( 3H]-5-HT), exposure to lysergic acid diethylamide or 5-methoxytryptamine decreased, in a concentration-dependent manner, the release of 3H-transmitter elicited by electrical stimulation. These inhibitory effects were antagonized by the 5-HT receptor antagonist methiothepin (1 microM). Exposure to methiothepin on its own increased in a concentration-dependent manner the electrically evoked overflow of [3H]-5-HT. Exposure to tricyclic antidepressants, like imipramine and amitriptyline, and to nontricyclic 5-HT uptake inhibitors, like paroxetine and citalopram, did not modify by themselves the electrically evoked overflow of [3H]-5-HT. Yet, the four inhibitors of neuronal uptake of 5-HT, antagonized the inhibition by lysergic acid diethylamide or 5-methoxytryptamine of the electrically induced release of [3H]-5-HT. After depletion of endogenous stores of 5-HT by pretreatment with para-chlorophenylalanine (300 mg/kg i.p.), the inhibitors of 5-HT uptake increased the electrically evoked release of [3H]-5-HT in a concentration-dependent manner. Their order of potency to enhance 5-HT overflow after pretreatment with parachlorophenylalanine paralleled their potency at inhibiting neuronal uptake of 5-HT (paroxetine = citalopram greater than imipramine greater than amitriptyline). In para-chlorophenylalanine-treated rat hypothalamic slices, these inhibitors of 5-HT uptake antagonized the inhibition by 5-HT autoreceptor agonists of the electrically evoked release of [3H]-5-HT to a similar extent than was observed in control rats. It is concluded that inhibition of 5-HT uptake reduces the effectiveness of 5-HT autoreceptor agonists to inhibit the electrically evoked release of [3H]-5-HT, irrespective of the chemical structure of the uptake inhibitor or of the levels of endogenous 5-HT achieved in the synaptic gap.

In rat hypothalamic slices prelabeled with [3H]-5-hydroxytryptamine ([3H]-5-HT), exposure to the 5-HT receptor agonist lysergic acid diethylamide (0.1-1 microM) or 5-methoxytryptamine (0.1-10 microM) decreased in a concentration-dependent manner the release of 3H-transmitter elicited by high K+ or electrical stimulation. Exposure to the 5-HT autoreceptor antagonist methiothepin (0.1-1 microM) increased in a concentration-dependent manner the K+ stimulation-evoked overflow of [3H]-5-HT and a similar increase was observed under conditions of electrical stimulation. In contrast, exposure to the nontricyclic 5-HT uptake inhibitor citalopram (0.1-1 microM) did not modify by itself the electrically evoked overflow of [3H]-5-HT, but increased in a concentration-dependent manner the release of 3H-transmitter elicited by K+ stimulation. This effect of citalopram on transmitter release was potentiated when the endogenous stores of 5-HT were depleted by pretreatment with para-chlorophenylalanine methyl ester (300 mg/kg i.p.). Citalopram was shown previously to antagonize the inhibition by lysergic acid diethylamide of the electrically evoked release of [3H]-5-HT in rat hypothalamic slices. Yet, this inhibitor of neuronal uptake of 5-HT did not antagonize the effects of lysergic acid diethylamide when the release of [3H]-5-HT was evoked by K+ depolarization. Electrical stimulation represents a more physiological experimental model for transmitter release than exposure to high K+, and therefore the interaction between 5-HT uptake blockade and presynaptic inhibitory 5-HT autoreceptors, observed in the hypothalamus with electrical stimulation but not with K+ depolarization, remains of biological relevance.

[3H]Imipramine and [3H]paroxetine label with high affinity a site associated with the serotonin transporter in brain and platelets. The maximum binding capacity (Bmax) of [3H]imipramine in platelets is reduced in untreated depressed patients, and it may represent a useful biological marker in depression. The existence of an endogenous ligand acting on the [3H]imipramine-recognition site to modulate the serotonin transporter has been proposed by several laboratories. 5-Methoxytryptoline inhibits [3H]imipramine binding and [3H]serotonin uptake in the nanomolar range. This compound has been reported to occur in the pineal gland, but probably only in trace amounts. While the physiological relevance of 5-methoxytryptoline or a close analogue remains an open question, the possibility exists that the 'endocoid' for the [3H]imipramine-recognition site plays a role in the pathogenesis of depression.