Relationship between serotonin and tryptamine receptors in the rat stomach fundus

Tryptamine and serotonin (5-HT) are relatively potent contractile agonists in the rat fundus, a tissue in which contraction to 5-HT is not mediated by interaction with 5-HT1 or 5-HT2 receptors. The identification of [3H]tryptamine binding sites in the brain and fundus that show high affinity for certain beta-carbolines raised the possibility that 5-HT and tryptamine may be interacting with a similar receptor that is best described as a tryptaminergic receptor in the fundus. The affinity of five 5-HT receptor antagonists, ketanserin, metergoline, 1-(1-naphthyl)piperazine, LY154930 and LY175041 was similar when 5-HT or tryptamine was the agonist, indicating that 5-HT and tryptamine are interacting with the same receptor in the fundus. Furthermore, maximum contractile response to both 5-HT and tryptamine was reduced to the same extent by the calcium channel blocker, diltiazem, and by the calmodulin inhibitor, trifluoperazine. Inasmuch as diltiazem and trifluoperazine did not similarly inhibit contraction to agents interacting with other receptors (i.e., carbamylcholine), these data are consistent with the contention that 5-HT and tryptamine are interacting with the same receptor in the fundus. Consistent with this conclusion is the observation that affinity of the beta-carbolines, harmaline and harmine was also similar when tryptamine or 5-HT was used as the agonist. However, affinity of the beta-carbolines for the tryptamine/5-HT receptor in the fundus was dramatically lower than reported for [3H]tryptamine binding sites in brain membranes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of contractions to serotonin, carbamylcholine and prostaglandin F2 alpha in rat stomach fundus

Contractile effects of serotonin were compared to those of carbamylcholine and prostaglandin (PG) F2 alpha in an effort to characterize serotonergic receptor activation in rat stomach fundus. All three agents elicited concentration-dependent contractions of fundal strips with serotonin (EC50 = 2 X 10(-10) M) being approximately 100-fold more potent than carbamylcholine (EC50 = 2 X 10(-8) M) and PGF2, (EC50 = 10(-8) M). The calcium channel blockers, diltiazem (5 X 10(-7) to 5 X 10(-5) M) and nitrendipine (10(-8) to 10(-5) M), attenuated responses markedly to serotonin and PGF2 alpha whereas having only minimal effects on carbamylcholine-induced contractions. Neither serotonin (10(-11) to 10(-5) M) nor PGF2 alpha (10(-9) to 10(-5) M) altered [3H]inositol monophosphate generation in fundus whereas carbamylcholine (10(-6) to 10(-5) M) increased significantly [3H]inositol monophosphate with 10(-5) M eliciting an 8-fold increase. Strips of fundus contracted submaximally with either serotonin, PGF2 alpha or carbamylcholine were compared for sensitivity to relaxants. Pinacidil (10(-8) to 10(-5) M) was equipotent (EC50 = 0.1 microM) in relaxing serotonin- and PGF2 alpha-contracted tissues. In contrast, pinacidil was 10-fold less potent in relaxing contractions produced by carbamylcholine. Likewise, nitroglycerin (10(-8) to 10(-4) M) and isoproterenol (10(-12) to 10(-7) M) were at least 10-fold more potent in relaxing serotonin- and PGF2 alpha- than carbamylcholine-induced contractions. Thus, in rat fundus, contractions associated with increased phosphoinositide hydrolysis may be more resistant to relaxation than are contractions not associated with altered phosphoinositide hydrolysis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Canine basilar artery contractions mediated by 5-hydroxytryptamine1A receptors

A series of 11 agents was analyzed at both 5-hydroxytryptamine1A (5-HT1A) sites labeled by [3H]-8-hydroxy-2-(N,N-dipropylamino)tetralin and total 5-HT1 binding sites labeled by [3H]-5-HT in rat brain membranes. Three distinct patterns of relative inhibition were noted. First, drugs such as 8-hydroxy-2-(N,N-dipropylamino)tetralin n-(3-acetylaminophenyl)piperazine hydrochloride, 2-[4-[4-(2-pyrimidinyl)-1-piperazinyl]-1,2-benziso-thiazol-3 -(2H)one-1,1-dioxidehydrochloride] and buspirone display 1.0 to 15 nM potency for the 5-HT1A subpopulation of 5-HT1 binding sites but are more than two orders of magnitude less potent at total 5-HT1 sites. Secondly, 5-methoxy-N,N-dimethyltryptamine and methysergide are approximately one order of magnitude more potent at 5-HT1A than total 5-HT1 sites. In the third group, 5-HT, d-lysergic acid diethylamide, 1-(m-trifluoromethylphenyl)piperazine, 5-methoxy-3-(1,2,3,6-tetrahydro-4-pyrimidinyl) 1H indole, quipazine and pirenperone are essentially equipotent at both 5-HT1A and 5-HT1 sites. Thus, the 5-HT1A binding site has a pharmacological profile which, depending on the agents studied, could be significantly different from the pharmacological profile derived from total 5-HT1 binding. Drug interactions were also analyzed with canine basilar artery segments using 5-HT, 10 putative serotonergic agonists and a selective 5-HT2 antagonist, pirenperone. The maximal contraction was obtained using 5-HT (Cmax = 6.6 +/- 0.6 g). However, each of the 10 other putative 5-HT agonists elicited a less forceful contraction of the canine basilar artery.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dissociation constants of serotonin agonists in the canine basilar artery correlate to Ki values at the 5-HT1A binding site

Attempts to correlate 5-hydroxytryptamine (5-HT) binding sites and the receptor mediating 5-HT-induced contractions in the canine basilar artery (CBA) have led to discrepant claims, particularly regarding the potencies of antagonists (which appear to act noncompetitively) in the CBA. In the classical theoretical model for noncompetitive antagonism, the antagonist does not inhibit agonist binding. Thus attempts to correlate noncompetitive antagonist pD2' values with binding measurements are inconsistent with this model and are of questionable significance. Only agonist dissociation constants (KA values) were measured in the CBA (by the irreversible antagonist method) and correlated to binding data. Logs of these KA values did not correlate with log Ki values for the inhibition of binding of the 5-HT2 ligand [3H] ketanserin (r = 0.2253, P greater than .05) or log IC50 values for inhibition of [3H]5-HT binding (r = 0.5732, P greater than 0.05), which labels both 5-HT1A and 5-HT1B sites. A correlation was observed between the CBA values and log Ki values for the 5-HT1A binding site subtype (r = 0.9456, P less than .01). The agonist ED50 and KA values were in most cases essentially identical, which suggests that in this tissue there are no spare receptors for 5-HT. Also, identical values were obtained when apparent noncompetitive antagonists were used in a manner analogous to the irreversible antagonist method to determine KA. These findings suggest the hypothesis that the apparent noncompetitive antagonism may be produced by slow dissociation of the antagonist-receptor complex, producing a pseudoirreversible action which is unmasked due to the lack of spare receptors.     

Characterization of 5-hydroxytryptamine receptors in rat stomach fundus

1-Arylpiperazines (MK-212, quipazine, m-chlorophenylpiperazine and m-trifluoromethylphenylpiperazine) caused a serotonin (5-HT) receptor-mediated contraction of rat fundic strips. m-chlorophenylpiperazine and m-trifluoromethylphenylpiperazine had high affinity for the receptor but little efficacy, whereas quipazine and MK-212 had lesser affinity and much greater efficacy. 5-HT itself was the most potent (EC50 = 6-9 nM) agonist and possessed the greatest affinity (KA = 9.7 nM). Assessment of receptor occupancy vs. functional response (as well as receptor alkylation studies) demonstrated a very small, if any, receptor reserve in this tissue. Several arylquinolizines were found to be competitive antagonists of 5-HT-induced contraction, the most potent being L-653,267 and rauwolscine (KB values = 1.9 and 3.8 nM). Clozapine, trazodone and propranolol were identified as less potent, competitive antagonists, whereas various ergolines (including LY 53857), L-646,462 (cyproheptadine analog) and mianserin were noncompetitive. Potent 5-HT2 receptor antagonists (pirenpirone and ketanserin) antagonized only weakly or were without effect against 5-HT, indicating that the fundic 5-HT receptor is not of the 5-HT2 subtype. Because the fundic receptor has high affinity for 5-HT (as does the 5-HT1 binding site in brain tissue), the possible correspondence of the fundic 5-HT receptor with the 5-HT1 recognition site in rat brain cortex was considered. 5-HT, the nonindole agonists (1-arylpiperazines) and the competitive antagonists all competed with [3H]-5-HT for the 5-HT1 site. However, all compounds except 5-HT had Hill slopes significantly less than 1.0, precluding a valid comparison with dissociation constants derived pharmacologically in the fundus. With respect to having a high affinity for 5-HT, the 5-HT receptor mediating contraction of fundic smooth muscle resembles the 5-HT1 recognition site (as defined in brain tissue by radioligand binding), but identity remains unproven.     

Improving the treatment of schizophrenia: focus on serotonin (5-HT)(1A) receptors

Although antagonism of mesolimbic dopamine D(2) receptors by neuroleptics such as haloperidol attenuates positive symptoms of schizophrenia, a significant population of "resistant" patients fails to respond while negative and cognitive symptoms are little modified. Furthermore, concomitant blockade of striatal D(2) receptors is associated with extrapyramidal motor side effects. The superior "atypical" antipsychotic profile of clozapine appears to reside in its broad pattern of interaction with D(2) receptors and a diversity of other monoaminergic sites. In this regard, serotonergic mechanisms are of particular relevance both in view of their modulation of dopaminergic transmission and their key role in the control of mood, cognition, and motor behavior. While most attention has focused on potential advantages of preferential 5-HT(2A) versus D(2) receptor blockade, 5-HT(1A) receptors likewise represent a valid target for improved antipsychotic agents. In this regard, rather than selective agents, ligands interacting with both 5-HT(1A) and D(2) receptors appear of interest. A modest level of efficacy appears optimal, that is, sufficient to engage highly sensitive 5-HT(1A) autoreceptors while blocking their low-sensitivity postsynaptic counterparts. Such a profile may counter negative and cognitive symptoms, improve mood, diminish extrapyramidal 5-HT(1A) motor side effects, and, perhaps, enhance efficacy in refractory patients. Notably, "partial agonist" properties of clozapine at 5-HT(1A) receptors may contribute to its distinctive functional profile. However, notwithstanding this compelling body of experimental data, clinical studies of antipsychotics interacting with 5-HT(1A) receptors are required to establish their genuine pertinence to the-hopefully improved-treatment of schizophrenia.     

Regulation of extracellular concentrations of 5-hydroxytryptamine (5-HT) in mouse striatum by 5-HT(1A) and 5-HT(1B) receptors

The ability of selective serotonin (5-HT) receptor agonists to reduce the extracellular concentration of 5-HT was examined in the striatum of awake, unrestrained mice by in vivo microdialysis. Systemic administration of either 8-OH-PIPAT (R-(+)-trans-8-hydroxy-2-[N-n-propyl-N-(3'-iodo-2'-propenyl)] aminotetralin), a novel 5-HT(1A) receptor agonist, or CP 94,253, a selective 5-HT(1B) receptor agonist, resulted in significant dose-related reductions of striatal 5-HT. The effect of 8-OH-PIPAT (1.0 mg/kg) was blocked by pretreatment with WAY 100635 (0.1 mg/kg), a selective 5-HT(1A) receptor antagonist, but it was not blocked by pretreatment with GR 127935 (0.056 mg/kg), a selective 5-HT(1B/1D) receptor antagonist. The effect of CP 94,253 (1.0 mg/kg) was blocked by pretreatment with GR 127935 (0.056 mg/kg) but was not blocked by pretreatment with WAY 100635 (0.1 mg/kg). Neither WAY 100635 nor GR 127935 altered extracellular 5-HT levels at the doses that were able to completely block the effects of either 8-OH-PIPAT or CP 94,253. The present findings suggest that, on systemic administration, both 8-OH-PIPAT and CP 94,253 are potent and selective agonists at the somatodendritic 5-HT(1A) autoreceptor and terminal 5-HT(1B/1D) autoreceptor, respectively, and are each able to cause decreases in extracellular levels of 5-HT in the mouse striatum by activating a distinct set of receptors.     

G-protein-linked serotonin receptors in mouse kidney exhibit identical properties to 5-HT1b receptors in brain

The serotonin 1b (5-HT1b) receptor is thought to mediate both pre- and postsynaptic actions of serotonin. Until recently 5-HT1b sites were thought to be present only in rodent brain. We now report the presence of high-affinity [125I]iodocyanopindolol [( 125I] ICYP) binding sites in the mouse renal medulla with properties identical to those of brain 5-HT1b receptors. In vitro receptor autoradiography demonstrates that [125I]ICYP binding is highly localized to the outer stripe of the renal medulla. Association and dissociation kinetics, saturation analysis and competition displacement analyses indicate that renal medullary [125I]ICYP binding sites exhibit identical properties with brain 5-HT1b receptors. Incubation of renal medullary or brain membranes with guanylimidodiphosphate results in a decreased affinity of 5-HT1b sites for 5-HT and [125I]ICYP; this can be reversed by the addition of a purified mixture of G proteins (Gi/Go). Treatment of brain or kidney membranes with N-ethylmaleimide results in a decrease in 5-HT1b binding which can also be restored by reconstitution with purified G proteins. Adenylyl cyclase from renal medullary homogenates or minces can be stimulated more than 3-fold by forskolin and attenuated by 5-HT. These results indicate that mouse kidney contains high-affinity 5-HT1b receptors with identical properties to those found in brain. These are localized in the outer stripe of the renal medulla and are functionally coupled to adenylyl cyclase inhibitor (Gi) G-proteins.     

Characterization of the human basilar artery contractile response to 5-HT and triptans

To study the contractile responses of the human basilar artery to 5-hydroxytryptamine (5-HT), sumatriptan, zolmitriptan and naratriptan, and to characterize the 5-HT receptor subtypes involved on those responses, human basilar artery rings were prepared for isometric contraction, protein isolation and Western blotting analysis. Concentration-response (CR) curves were made for all agonists in the absence or in the presence of selective antagonists at 5-HT1B (cyanopindolol), 5-HT1D (BRL 15,572) and 5-HT2 (ketanserin) receptors. We also used anti-5-HT1B and 5-HT1D receptor antibodies to search for the expression of protein of these receptor subtypes. From the CR curves, the relative intrinsic activity and potency of these agonists were determined. The ranking order for the intrinsic activity was 5-HT > or = sumatriptan > zolmitriptan > or = naratriptan, whereas that for the potency was zolmitriptan > or = 5-HT > or = sumatriptan > naratriptan. Our results also show that the human basilar artery seems to have a mixed population of 5-HT1B/1D receptors mediating the contractile response to triptans, which is also suggested by the expression of both receptor subtypes. There is also a population of 5-HT2 receptors for which the antimigraine drugs used have no apparent affinity. From this study, one can conclude that the second generation triptans have lower contractile capacity than sumatriptan, suggesting that they have a better cerebrovascular safety profile.     

Role of 5-HT2 receptors in serotonin-induced contractions of nonvascular smooth muscle

Vascular receptors responsible for serotonin-induced contractions are of the 5-HT2 subtype (site in brain cortical membranes that is preferentially radiolabeled by [3H]spiperone) whereas serotonin receptors mediating contraction in nonvascular smooth muscle have not been extensively studied. The present in vitro studies using the 5-HT2 receptor antagonists ketanserin, LY53857 and 1-(1-naphthyl)piperazine show that serotonin-induced contractions in the rat uterus and guinea-pig trachea are also mediated by interaction with 5-HT2 receptors. Prazosin, but not the serotonin receptor antagonists, blocked serotonin-induced contractions in the rat vas deferens, indicating that alpha adrenergic and not 5-HT1 or 5-HT2 receptors mediate the contractile response to serotonin in this tissue. Because selective 5-HT2 receptor antagonists did not block contractions to serotonin in the rat fundus or guinea-pig ileum, receptors in these gastrointestinal tissues are clearly not 5-HT2. However, contractions to serotonin in the fundus but not in the ileum were blocked by certain antagonists [metergoline and 1-(1-naphthyl)piperazine] demonstrating that the receptors involved in serotonin-induced contractions in the fundus are different from the ileum. Other differences between the fundus and ileum in serotonin-induced contractions include: 1) the potency of serotonin is greater in the fundus than in the ileum; and 2) the primary action of serotonin in the fundus is activation of a postsynaptic receptor on the smooth muscle whereas, in the ileum, serotonin exerts an indirect neuronal action to effect acetylcholine release.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hyperlocomotive and discriminative stimulus effects of cocaine are under the control of serotonin(2C) (5-HT(2C)) receptors in rat prefrontal cortex

The serotonin2C (5-hydroxytryptamine2C; 5-HT2C) receptor (5-HT2CR) is found in abundance in dopamine (DA) mesocorticolimbic pathways and is one of the important target proteins that modulates the behavioral effects of cocaine. In the present study, the hypothesis was tested that the 5-HT2CR in the prefrontal cortex (PFC) may control either spontaneous or cocaine-evoked locomotor activity as well as the discriminative stimulus properties of cocaine. In male Sprague-Dawley rats implanted with bilateral cannulae aimed at the PFC, local microinjections of the preferential 5-HT2CR agonist 6-chloro-2-(1-piperazinyl)pyrazine hydrochloride (MK 212) (0.05-0.5 microg/side) did not alter spontaneous activity, but dose-dependently decreased horizontal hyperactivity evoked by cocaine (10 mg/kg i.p.). Given alone, the selective 5-HT2CR antagonist 8-[5-(2,4-dimethoxy-5-(4-trifluorophenylsulfonamido)phenyl-5-oxopentyl]-1,3,8-triazo-spiro[4.5]decane-2,4-dione hydrochloride (RS 102221) (5 microg/side) increased basal locomotor activity of rats expressed in the vertical plane. Microinjections of RS 102221 (5 microg/side, but not 0.15-1.5 microg/side) significantly enhanced the horizontal activity induced by cocaine (10 mg/kg). In rats trained to discriminate cocaine (10 mg/kg i.p.) from saline (i.p.) in a two-lever, water-reinforced fixed ratio 20 task, intra-PFC microinjections of MK 212 (0.05 and 0.5 microg/side) did not substitute for cocaine, but attenuated the stimulus effects of cocaine. However, intra-PFC microinjections of RS 102221 (1.5 and 5 microg/side) evoked 13 and 40% cocaine-lever responding when tested alone and enhanced the recognition of cocaine. These data indicate that the PFC is a brain site at which the 5-HT2CR exerts an inhibitory control over the hyperactive and discriminative stimulus effects of cocaine known to be dependent upon activation of the DA mesoaccumbens circuit.     

Spinal 5-HT(2) and 5-HT(3) receptors mediate low, but not high, frequency TENS-induced antihyperalgesia in rats

Transcutaneous electrical nerve stimulation (TENS) is a form of non-pharmacological treatment for pain. Involvement of descending inhibitory systems is implicated in TENS-induced analgesia. In the present study, the roles of spinal 5-HT and alpha(2)-adrenoceptors in TENS analgesia were investigated in rats. Hyperalgesia was induced by inflaming the knee joint with 3% kaolin-carrageenan mixture and assessed by measuring paw withdrawal latency (PWL) to heat before and 4 h after injection. The (1). alpha(2)-adrenergic antagonist yohimbine (30 microg), (2). 5-HT antagonist methysergide (5-HT(1). and 5-HT(2). 30 microg), one of the 5-HT receptor subtype antagonists, (3). NAN-190 (5-HT(1A), 15 microg), (4). ketanserin (5-HT(2A), 30 microg), (5). MDL-72222 (5-HT(3), 12 microg), or (6). vehicle was administered intrathecally prior to TENS treatment. Low (4 Hz) or high (100 Hz) frequency TENS at sensory intensity was then applied to the inflamed knee for 20 min and PWL was determined. Selectivity of the antagonists used was confirmed using respective agonists administered intrathecally. Yohimbine had no effect on the antihyperalgesia produced by low or high frequency TENS. Methysergide and MDL-72222 prevented the antihyperalgesia produced by low, but not high, frequency TENS. Ketanserin attenuated the antihyperalgesic effects of low frequency TENS whereas NAN-190 had no effect. The results from the present study show that spinal 5-HT receptors mediate low, but not high, frequency TENS-induced antihyperalgesia through activation of 5-HT(2A) and 5-HT(3) receptors in rats. Furthermore, spinal noradrenergic receptors are not involved in either low or high frequency TENS antihyperalgesia.     

Activation of serotonin2 (5-HT2) receptors by quipazine increases arterial pressure and renin secretion in conscious rats

Quipazine, a nonselective serotonin (5-HT) agonist, has been shown to increase plasma renin activity (PRA). The present study examined the effects of quipazine on mean arterial pressure (MAP), heart rate (HR) and PRA in conscious, chronically catheterized male rats. Quipazine caused dose (0.3-3.0 mg/kg i.v.)-and time (up to 30 min)-dependent increases in MAP and PRA. The maximum increases in MAP (control = 94 +/- 2 mm Hg, 3 mg/kg = 155 +/- 1 mm Hg) and PRA (nanograms of angiotensin 1 per milliliter per hour; control = 2.5 +/- 0.2, 3 mg/kg = 25.2 +/- 5.9) were observed 5 min after quipazine. HR tended to decrease, but a significant bradycardia was observed only 15 min after 3 mg/kg. The selective 5-HT2 antagonist LY 53857 (1 mg/kg i.v.) did not affect MAP, HR or PRA per se, but at 0.03 to 1.0 mg/kg totally abolished the pressor response to quipazine (3 mg/kg). At 0.01 mg/kg, LY 53857 attenuated quipazine-induced hypertension, whereas 0.003 mg/kg was ineffective. Total blockade of quipazine-induced renin secretion was produced by LY 53857 at 0.003 mg/kg, and the response was still reduced by 50% at 0.001 mg/kg. In summary, although quipazine increases arterial pressure and renin secretion, endogenous 5-HT does not tonically control MAP or PRA in conscious, unrestrained, normotensive rats through 5-HT2 receptors. The 10-fold difference in the dose of LY 53857 necessary to block the pressor and renin responses may be due to subtle differences in receptor subtypes, or to pharmacokinetic properties favoring antagonism of quipazine-induced renin secretion.     

Temporal regulation of agonist efficacy at 5-hydroxytryptamine (5-HT)1A and 5-HT 1B receptors

Coactivation of purinergic (P 2Y) receptors reduces agonist efficacy at serotonin 1B (5-HT 1B), but not 5-HT 1A receptors. Herein, we report that pretreatment for 5 min with the P 2Y receptor agonist ATP reduced agonist responsiveness at the 5-HT 1A, but not at the 5-HT 1B, receptor. The effect of ATP pretreatment on the 5-HT 1A receptor response rapidly reversed within a 10 min time frame between P 2Y receptor and 5-HT 1A receptor activation. ATP pretreatment effects on 5-HT 1A agonist responsiveness were blocked by the protein kinase inhibitors staurosporine and bisindolylmaleimide, suggesting that the ATP-mediated temporal regulation involves activation of protein kinase C (PKC). Moreover, the temporal effect of ATP was blocked by incubation with 1% ethanol, suggesting that consequences of phospholipase D (PLD) activation play a role. ATP pretreatment blocked the inhibitory effect produced by 5-HT 2C receptor activation on the 5-HT 1A, but not the 5-HT 1B, receptor response, suggesting that the 5-HT 1A receptor itself was the target for PLD/PKC action. Finally, ethanol did not block the reduction in responsiveness of the 5-HT 1A receptor system produced by activation of PKC with phorbol ester treatment, suggesting that PKC activation lies downstream of PLD. Taken together, these data suggest that activation of P 2Y receptors can reduce responsiveness of the 5-HT 1A receptor system via a PLD/PKC-dependent mechanism that is highly dependent upon the temporal pattern of receptor activation. Moreover, this work underscores the importance of time as a variable in receptor signaling cross talk and serves to further illustrate differences between the 5-HT 1A and 5-HT 1B receptor systems.     

Effect of nitrendipine, diltiazem, trifluoperazine and pimozide on serotonin2 (5-HT2) receptor activation in the rat uterus and jugular vein

The present study explored the calcium source used in serotonin (5-HT)-induced contractions mediated by 5-HT2 receptor activation in the rat uterus and jugular vein in vitro. In the rat uterus, the calcium channel antagonists, nitrendipine and diltiazem, and the neuroleptic agents, trifluoperazine (TFP) and pimozide, antagonized potently and noncompetitively 5-HT-induced contractions. These data are compatible with the contention that 5-HT-induced contractions in uterine smooth muscle require extracellular sources of calcium. In contrast, neither diltiazem nor nitrendipine inhibited the contractile response to 5-HT in the rat jugular vein although 5-HT-induced contractions in the jugular vein required the presence of extracellular calcium. This difference in sensitivity to calcium channel antagonists between the uterus and jugular vein was not related to differences in the receptor occupancy-response curves for 5-HT as these were similar in the rat uterus and jugular vein. Furthermore, in the jugular vein, neither diltiazem nor nitrendipine were markedly effective in blocking contractions produced by potassium chloride, an agent that activates voltage-dependent calcium channels, suggesting that this tissue lacks calcium channels susceptible to blockade by conventional calcium channel antagonists. Although responses to 5-HT in the jugular vein were only affected marginally by calcium channel antagonists, TFP and pimozide produced concentration-dependent rightward parallel shifts of 5-HT-induced contractions in the jugular vein. Furthermore, both TFP and pimozide showed high affinity at 5-HT2 binding sites in rat brain cortical membranes and the apparent dissociation constant at 5-HT2 receptors in the jugular vein was 38 and 31 nM, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Desensitization of 5-HT1A receptors by 5-HT2A receptors in neuroendocrine neurons in vivo

An imbalance between serotonin-2A (5-HT2A) and 5-HT1A receptors may underlie several mood disorders. The present studies determined whether 5-HT2A receptors interact with 5-HT1A receptors in the rat hypothalamic paraventricular nucleus (PVN). The sensitivity of the hypothalamic 5-HT1A receptors was measured as oxytocin and adrenocorticotropic hormone (ACTH) responses to the 5-HT1A receptor agonist (+)-8-hydroxy-2-(di-n-propylamino) tetralin hydrobromide [(+)8-OH-DPAT] (40 microg/kg s.c.). The 5-HT(2A/2C) receptor agonist (-)DOI [(-)-1-(2,5-dimethoxy-4-iodophenyl)2-aminopropane HCl] (1 mg/kg s.c.) injected 2 h prior to (+)8-OH-DPAT significantly reduced the oxytocin and ACTH responses to (+)8-OH-DPAT, producing a heterologous desensitization of the 5-HT1A receptors. Microinjection of the 5-HT2A receptor antagonist MDL100,907 [(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenylethyl)]-4-piperidinemethanol; 0, 10, or 20 nmol, 15 min prior to (-)DOI] into the PVN dose-dependently prevented the desensitization of 5-HT1A receptors induced by the 5-HT2A receptor agonist (-)DOI. Double-label immunocytochemistry revealed a high degree of colocalization of 5-HT1A and 5-HT2A receptors in the oxytocin and corticotropin-releasing factor neurons of the PVN. Thus, activation of 5-HT2A receptors in the PVN may directly induce a heterologous desensitization of 5-HT1A receptors within individual neuroendocrine cells. These findings may provide insight into the long-term adaptation of 5-HT1A receptor signaling after changes in function of 5-HT2A receptors; for example, during pharmacotherapy of mood disorders.     

Effects of serotonin (5-hydroxytryptamine, 5-HT) reuptake inhibition plus 5-HT(2A) receptor antagonism on the firing activity of norepinephrine neurons

YM992 [(S)-2-[[(7-fluoro-4-indanyl)oxy]methyl]morpholine monohydrochloride] is a selective serotonin (5-hydroxytryptamine; 5-HT) reuptake inhibitor (SSRI) and a potent 5-HT(2A) antagonist. The aim of the present study was to assess, using in vivo extracellular unitary recordings, the effect of acute and sustained administration of YM992 (40 mg kg(-1) day(-1) s.c., using osmotic minipumps) on the spontaneous firing activity of locus coeruleus (LC) norepinephrine (NE) neurons. Acute intravenous injection of YM992 (4 mg kg(-1)) significantly decreased NE neuron firing activity by 29% and blocked the inhibitory effect of a subsequent injection of the 5-HT(2) agonist DOI [1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride]. A 2-day treatment with YM992 decreased the firing rate of NE neurons by 66%, whereas a partial recovery was observed after a 7-day treatment and a complete one after a 21-day treatment. Following the injection of the alpha(2)-adrenoceptor antagonist idazoxan (1 mg kg(-1) i.v.), NE neuron firing was equalized in controls and 2-day YM992-treated rats. This put into evidence an increased degree of activation of alpha(2)-adrenergic autoreceptors in the treated rats. The suppressant effect of the alpha(2)-adrenoceptor agonist clonidine was significantly decreased in long-term YM992-treated rats. The recovery of LC firing activity after long-term YM992 administration could thus be explained by a decreased sensitivity of alpha(2)-adrenergic autoreceptors. Sustained SSRI administration leads to a gradual reduction of the firing activity of NE neurons during long-term administration, whereas YM992 produced opposite effects. The exact basis for the increased synaptic availability of NE by YM992 remains to be elucidated. This NE activity, resulting from 5-HT reuptake inhibition plus 5-HT(2A) receptor antagonism, might confer additional benefits in affective and anxiety disorders.     

Brain 5-HT1 and 5-HT2 binding sites following portacaval shunt in the rat

Brain serotonin 5-HT1 and 5-HT2 binding properties were investigated in experimental chronic portal-systemic encephalopathy (PSE). End-to-side portacaval shunted (PCS) rats were subjected to open field behavioral testing (spontaneous activity and exploration) 3 weeks after the shunt procedure. Each individual animal was then assayed for 5-HT1 and 5-HT2 binding properties (Bmax and KD) in the cortex + hippocampus by the use of radioligand binding and rapid filtration technique. (3H)serotonin was used to label 5-HT1 binding sites and (3H)ketanserin to label 5-HT2 binding sites. Results revealed that the PCS rats exhibited significant behavioral changes with decreased spontaneous activity and exploratory behavior as compared with sham-operated controls (sham). The affinity for, and the number of, 5-HT1 and 5-HT2 binding sites, respectively, were not different between PCS and sham rats. The brain 5-HT1 and 5-HT2 binding properties were within the range of Bmax and KD previously reported for normal rats when similar techniques are used. This first report in PCS rats on the subject of brain 5-HT1 and 5-HT2 binding properties demonstrates that no major alterations are likely to occur. This contrasts the knowledge of a markedly increased brain serotonin synthesis rate in the PCS rat, suggesting minor functional relevance of the perturbed brain serotonin metabolism associated with chronic PSE.     

Characterization of sumatriptan-induced contractions in human isolated blood vessels using selective 5-HT(1B) and 5-HT(1D) receptor antagonists and in situ hybridization

The 5-HT(1B/1D) receptor agonist sumatriptan is effective in aborting acute attacks of migraine and is known to cause constriction of cranial arteries as well as some peripheral blood vessels. The present study set out to investigate whether 5-HT(1B) and/or 5-HT(1D) receptors mediate contractions of the human isolated middle meningeal and temporal arteries (models for anti-migraine efficacy) and coronary artery and saphenous vein (models for side-effect potential). Concentration-response curves were made with sumatriptan (1 nm-100 microm) in blood vessels in the absence or presence of selective antagonists at 5-HT(1B) (SB224289) and 5-HT(1D) (BRL15572) receptors. SB224289 antagonized sumatriptan-induced contractions in all blood vessels, although the antagonism profile was different amongst these blood vessels. In the temporal artery, SB224289 abolished contraction to sumatriptan, whereas in the middle meningeal artery and saphenous vein sumatriptan-induced contractions were blocked in an insurmountable fashion. Moreover, SB224289 acted as a weak surmountable antagonist in the coronary artery (pK(B): 6.4 +/- 0.2). In contrast, BRL15572 had little or no effect on sumatriptan-induced contractions in the four blood vessels investigated. In situ hybridization revealed the expression of 5-HT(1B) receptor mRNA in the smooth muscle as well as endothelial cells of the blood vessels, whereas the mRNA for the 5-HT(1D) receptor was only very weakly expressed. These results show that the 5-HT(1B) receptor is primarily involved in sumatriptan-induced contractions of human cranial as well as peripheral blood vessels.     

5-Hydroxytryptamine (5-HT)-induced endothelium-dependent relaxation of pig coronary arteries is mediated by 5-HT receptors similar to the 5-HT1D receptor subtype

The 5-hydroxytryptamine (5-HT) receptor mediating endothelium-dependent relaxation of pig coronary arteries was characterized using a variety of 5-HT receptor agonists and antagonists. Unrubbed (with endothelium preserved) rings precontracted by prostaglandin F2 alpha in the presence of ketanserin relaxed in an endothelium-dependent manner to 5-HT, 5-carboxamidotryptamine and 5-methoxytryptamine with about equal potency and efficacy. By comparison, bufotenine, 3-(dimethylamino)ethyl-N-methyl-1H-indole-5-methane sulfonamide, (-)-alpha-methyl-5-HT,N,N-dipropyl-5-carboxamidotryptamine and 5-methoxy-3-(1,2,3,6-tetrahydro-4-pyridinyl)-1H indole were half-efficient and other drugs [in particular the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin] were inactive as agonists up to 0.1 mM. The effect of 5-carboxamidotryptamine was antagonized in an apparently competitive manner by 15 drugs. Among the most potent antagonists (mean pKB value) were the nonselective 5-HT receptor antagonists, methiothepin (7.30) and metergoline (6.86), the 5-HT1A/5-HT1D receptor ligand, 1-[2-(4-amino-phenyl)ethyl]-4-(3-trifluoromethylphenyl)-piperazine (7.02), the 5-HT1A/5-HT1B/5-HT1D receptor ligand, 7-trifluoromethyl-4-(4-methyl-1-piperazinyl)-pyrrolo[1,2,-a]quinoxaline 1 (6.73) and yohimbine (6.37). Selective ligands for 5-HT1A receptors were either inactive [8-hydroxy-2-(di-n-propylamino)tetralin hydrobromide] or poorly active (spiperone, 4.44). Beta-adrenoceptor antagonists with affinity for 5-HT1A and 5-HT1B receptors weakly antagonized the effect of 5-carboxamidotryptamine (pKB values less than or equal to 5.32), as did the 5-HT1c/5-HT2 receptor antagonist, mesulergine (5.30) and the yohimbine isomer, corynanthine (4.85). Methysergide was clearly a noncompetitive antagonist.(ABSTRACT TRUNCATED AT 250 WORDS)