Serotonin Inhibition of the NMDA Receptor/Nitric Oxide/Cyclic GMP Pathway in Rat Cerebellum: Involvement of 5-Hydroxytryptamine2C Receptors

Abstract: Previous studies have shown that 5-hydroxytryptamine (5-HT) can potently inhibit glutamatergic transmission in rat cerebellum through the activation of multiple 5-HT receptors. The aim of this study was to subclassify the 5-HT₂ receptor mediating inhibition of the cyclic GMP response elicited by N -methyl- d -aspartate in adult rat cerebellar slices. Seven receptor antagonists, endowed with relative selectivities for the 5-HT_2A, 5-HT_2B, and 5-HT_2C subtypes, differentially affected the inhibition by (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane of the cyclic GMP response, suggesting that the receptor involved belongs to the 5-HT_2C subtype.     

Mianserin-Induced Down-Regulation of Human 5-Hydroxytryptamine2A and 5-Hydroxytryptamine2C Receptors Stably Expressed in the Human Neuroblastoma Cell Line SH-SY5Y

Abstract: We have assessed the ability of the serotonergic antagonist mianserin to modulate the number and functional activity of human 5-hydroxytryptamine_2A (5-HT_2A) and 5-HT_2C receptors stably expressed in the human neuroblastoma cell line SH-SY5Y. Incubation of cells expressing the 5-HT_2A receptor with mianserin (100 n M ) for 24 h caused a significant decrease (48%) in the binding capacity of [³H]ketanserin. This receptor down-regulation was associated with a corresponding decrease in the maximal production of inositol phosphates induced by 5-HT but not by carbachol. Exposure of cells expressing the 5-HT_2C receptor to mianserin (100 n M ) for 72 h but not for 24 h similarly resulted in a significant reduction (44%) in [³H]mesulergine binding. Corresponding analysis of inositol phosphate production by 5-HT at the 5-HT_2C receptor after incubation with mianserin showed no change in maximal response after 24 h. No change in the binding capacity of either radioligand was seen after incubation with mianserin for 1 h. A decrease in the binding affinity of both radioligands was also observed after mianserin treatment, but this decrease was similar after 1 h of incubation to that seen after 24 or 72 h, and was probably due to the retention of mianserin within the tissue. We conclude that antagonist down-regulation is evident at human 5-HT_2A and 5-HT_2C receptors stably expressed in a human neuroblastoma cell line and is probably mediated by a direct action of mianserin at the receptor.     

Endogenous serotonin and serotonin2C receptors are involved in the ability of M100907 to suppress cortical glutamate release induced by NMDA receptor blockade

Blockade of NMDA receptors by intracortical infusion of 3-( R )-2-carboxypiperazin-4-propyl-1-phosphonic acid (CPP) increases glutamate (GLU) and serotonin (5-HT) release in the medial prefrontal cortex and impairs attentional performance in the 5-choice serial reaction time task. These effects are prevented by the 5-HT_2A receptor antagonist, ( R )-(+)-(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenyl)ethyl]-4-piperidine methanol (M100907). We explored the roles of endogenous 5-HT and 5-HT_1A and 5-HT_2C receptors in the mechanisms by which M100907 suppresses CPP-induced release of cortical GLU and 5-HT using in vivo microdialysis. CPP raised extracellular GLU and 5-HT by about 250% and 170% respectively. The 5-HT synthesis inhibitor, p -chlorophenylalanine (300 mg/kg), prevented M100907 suppressing CPP-induced GLU release. The effect of M100907 on these rises of GLU and 5-HT and attentional performance deficit was mimicked by the 5-HT_2C receptor agonist, ( S )-2-(6-chloro-5-fluoroindol-1-yl)-1-methylethylamine fumarate, (Ro60-0175, 30 μg/kg) while intra-mPFC (SB242084, 6-chloro-5-methyl-1-[[2-[(2-methyl-3-pyridyl)oxy]-5-pyridyl]carbamoyl]-indoline, 0.1 μM), a 5-HT_2C receptor antagonist, prevented the effect of M100907 on extracellular GLU. The 5-HT_1A receptor antagonist, N -[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]- N -(2-pyridinyl)cyclohexane carboxenide trihydrochloride (100 μM) abolished the effect of M100907 on the CPP-induced 5-HT release. The data show that blockade of 5-HT_2A receptors is not sufficient to suppress the CPP-induced rise of extracellular GLU and 5-HT and suggest that M100907 suppresses GLU release induced by CPP by enhancing the action of endogenous 5-HT on 5-HT_2C receptors.     

CB1 knockout mice display impaired functionality of 5-HT1A and 5-HT2A/C receptors

Interaction between brain endocannabinoid (EC) and serotonin (5-HT) systems was investigated by examining 5-HT-dependent behavioral and biochemical responses in CB₁ receptor knockout mice. CB₁ knockout animals exhibited a significant reduction in the induction of head twitches and paw tremor by the 5-HT_2A/C receptor selective agonist (±) DOI, as well as a reduced hypothermic response following administration of the 5-HT_1A receptor agonist (±)-8-OH-DPAT. Additionally, exposure to the tail suspension test induced enhanced despair responses in CB₁ knockout mice. However, the tricyclic antidepressant imipramine and the 5-HT selective reuptake inhibitor fluoxetine induced similar decreases in the time of immobility in the tail suspension test in CB₁ receptor knockout and wild-type mice. No differences were found between both genotypes with regard to 5-HT_2A receptor and 5-HT_1A receptors levels, measured by autoradiography in different brain areas. However, a significant decrease in the ability of both, the 5-HT_1A receptor agonist (±)-8-OH-DPAT and the 5-HT_2A/C receptor agonist (−)DOI, to stimulate [³⁵S]GTPγS binding was detected in the hippocampal CA₁ area and fronto-parietal cortex of CB₁ receptor knockout mice, respectively. This study provides evidence that CB₁ receptors are involved in the regulation of serotonergic responses mediated by 5-HT_2A/C and 5-HT_1A receptors, and suggests that a reduced coupling of 5-HT_1A and 5-HT_2A receptors to G proteins might be involved in these effects.     

Primary Structure of the Human Platelet Serotonin 5-HT2A Receptor: Identity with Frontal Cortex Serotonin 5-HT2A Receptor

Abstract: Previous radioligand binding studies have demonstrated human platelet serotonin_2A (5-HT_2A) receptor binding sites. Pharmacological similarities between platelet and frontal cortex 5-HT_2A receptor binding parameters have been demonstrated. However, it is not clear whether the platelet 5-HT_2A receptor primary structure is identical to that of the brain receptor. Three overlapping cDNAs were obtained to span completely the coding region of the 5-HT_2A receptor. These clones were sequenced with external and internal primers. The nucleotide sequence of human platelet 5-HT_2A cDNA was identical to that reported for the human frontal cortex 5-HT_2A receptor, except for nucleotide 102 (T → C), which has been reported to represent a normal DNA polymorphism that does not alter the amino acid sequence. This finding may have implications in the study of neuropsychiatric disorders for which altered platelet 5-HT_2A receptor binding has been demonstrated.     

Antisense Oligonucleotide-Induced Reduction in 5-Hydroxytryptamine7 Receptors in the Rat Hypothalamus Without Alteration in Exploratory Behaviour or Neuroendocrine Function

Abstract: The effect of a 5-hydroxytryptamine₇ (5-HT₇) receptor-directed antisense oligonucleotide on rat behaviour and neuroendocrine function was investigated. Six days of intracerebroventricular 5-HT₇ antisense oligonucleotide treatment significantly reduced [³H]5-HT binding to hypothalamic 5-HT₇ receptors, whereas cortical 5-HT_2C density remained unchanged. In rats on a food-restricted diet, both antisense and mismatch oligonucleotides reduced food intake and body weight compared with that in vehicle-treated controls by day 4 of administration. 5-HT₇ antisense oligonucleotide administration did not affect exploratory or locomotor activity in photocell activity monitors on day 4 or elevated plus-maze behaviour on day 6 of intracerebroventricular treatment. 5-HT₇ antisense oligonucleotide did not affect plasma corticosterone or prolactin levels or 5-HT turnover in either 5-HT cell body or terminal areas. These data demonstrate that intracerebroventricular 5-HT₇ antisense oligonucleotide administration selectively reduced rat hypothalamic 5-HT₇ receptor density without affecting any of the biochemical or behavioural measures. The results suggest that this antisense protocol could be a valuable tool to investigate central 5-HT₇ receptor functions, and that this receptor is not critical for the control of neuroendocrine function or food intake.     

Activating Mutations of the Serotonin 5-HT2C Receptor

Abstract: Site-directed mutagenesis was performed to create a mutant serotonin 5-HT_2C receptor that would mimic the active conformation of the native receptor. Structural alteration of receptor conformation was achieved by changing amino acid no. 312 from serine to phenylalanine (S312F) or lysine (S312K). After expression in COS-7 cells, the binding affinity of 5-HT for [³H]-mesulergine-labeled 5-HT_2C receptors increased from 203 n M (native) to 76 n M for S312F and 6.6 n M for S312K mutant receptors. 5-HT potency for stimulation of phosphatidylinositol (PI) hydrolysis increased from 70 n M (native) to 28 n M for S312F and 2.7 n M for S312K mutant receptors. The mutant receptors were constitutively active, stimulating PI hydrolysis in the absence of agonist. S312F and S312K mutations resulted in twofold and five-fold increases, respectively, in basal levels of PI hydrolysis. Mianserin and mesulergine displayed inverse agonist activity by decreasing basal levels of PI hydrolysis stimulated by S312K mutant receptors. [³H]5-HT and [³H]-mesulergine labeled the same number of S312K mutant receptors and 5'-guanylylimidodiphosphate had no effect on [³H]5-HT binding. These results indicate that serine → lysine mutation at amino acid no. 312 produces an agonist high-affinity state of the 5-HT_2C receptor that spontaneously couples to G proteins and stimulates PI hydrolysis in the absence of agonist.     

In vivo evidence that constitutive activity of serotonin2C receptors in the medial prefrontal cortex participates in the control of dopamine release in the rat nucleus accumbens: differential effects of inverse agonist versus antagonist

Control of the mesoaccumbens dopamine (DA) pathway by central serotonin_2C receptors (5-HT_2CRs) involves different 5-HT_2CR populations located within multiple brain areas. Here, using in vivo microdialysis in halothane-anesthetized rats, we assessed the role of medial prefrontal cortex (mPFC) 5-HT_2CRs in the control of basal and activated accumbal DA outflow, to identify the modalities of their recruitment and the role of 5-HT_2CR constitutive activity. Intra-mPFC injection of the 5-HT_2CR inverse agonist SB 206553 (0.5 μg/0.2 μL), without effect by itself, decreased accumbal DA outflow induced by morphine (2.5–10 mg/kg, s.c.), haloperidol (0.01 mg/kg, s.c.) or GBR 12909 (2.5 mg/kg, i.p.). Conversely, intra-mPFC injection of the 5-HT_2CR antagonist SB 242084 (0.5 μg/0.2 μL), without effect by itself, decreased the effect of 10 mg/kg morphine, the only drug enhancing basal 5-HT outflow in the mPFC. The inhibitory effect of SB 206553 on 2.5 mg/kg morphine-stimulated DA outflow was suppressed by the concomitant intra-mPFC injection of SB 242084. Finally, changes of basal DA outflow induced by the 5-HT_2CR agonist Ro 60-0175 (3 mg/kg, i.p.) or SB 206553 (5 mg/kg, i.p.) were unaffected by intra-mPFC injection of SB 242084. These results, showing that 5-HT_2CR antagonist and inverse agonist behave differently in vivo, demonstrate that mPFC 5-HT_2CRs facilitate activated accumbal DA outflow and that 5-HT_2CR constitutive activity participates in this interaction.     

Cloning, Characterization, and Chromosomal Localization of a Human 5-HT6 Serotonin Receptor

Abstract: We describe the cloning and characterization of a human 5-HT₆ serotonin receptor. The open reading frame is interrupted by two introns in positions corresponding to the third cytoplasmic loop and the third extracellular loop. The human 5-HT₆ cDNA encodes a 440-amino-acid polypeptide whose sequence diverges significantly from that published for the rat 5-HT₆ receptor. Resequencing of the rat cDNA revealed a sequencing error producing a frame shift within the open reading frame. The human 5-HT₆ amino acid sequence is 89% similar to the corrected rat sequence. The recombinant human 5-HT₆ receptor is positively coupled to adenylyl cyclase and has pharmacological properties similar to the rat receptor with high affinity for several typical and atypical antipsychotics, including clozapine. The receptor is expressed in several human brain regions, most prominently in the caudate nucleus. The gene for the receptor maps to the human chromosome region 1p35–p36. This localization overlaps that established for the serotonin 5-HT_1Dα receptor, suggesting that these may be closely linked. Comparison of genomic and cDNA clones for the human 5-HT₆ receptor also reveals an Rsa I restriction fragment length polymorphism within the coding region.     

The brain 5-HT4 receptor binding is down-regulated in the Flinders Sensitive Line depression model and in response to paroxetine administration

The 5-hydroxytryptamine (5-HT₄) receptor may be implicated in depression and is a new potential target for antidepressant treatment. We have investigated the brain 5-HT₄ receptor [³H]SB207145 binding in the Flinders Sensitive Line rat depression model by quantitative receptor autoradiography, and related this to 5-HT transporter ( S )-[ N -methyl-³H]citalopram binding. We also determined the regulation of 5-HT₄ receptor binding by 1, 14, and 21 days of paroxetine administration and subchronic 5-HT depletion, and compared this with changes in 5-HT_2A receptor [³H]MDL100907 binding. In the Flinders Sensitive Line, the 5-HT₄ receptor and 5-HT transporter binding were decreased in the dorsal and ventral hippocampus, and the changes in binding were directly correlated within the dorsal hippocampus. Chronic but not acute paroxetine administration caused a 16–47% down-regulation of 5-HT₄ receptor binding in all regions evaluated including the basal ganglia and hippocampus, while 5-HT depletion increased the 5-HT₄ receptor binding in the dorsal hippocampus, hypothalamus, and lateral globus pallidus. In comparison, the 5-HT_2A receptor binding was decreased in the frontal and cingulate cortices after chronic paroxetine administration, and markedly reduced in several regions after 5-HT depletion. Thus, the 5-HT₄ receptor binding was decreased in the Flinders Sensitive Line depression model and in response to chronic paroxetine administration.     

Native Serotonin 5-HT3 Receptors Expressed in Xenopus Oocytes Differ from Homopentameric 5-HT3 Receptors

Abstract: Efficacies of the 5-hydroxytryptamine (serotonin) 5-HT₃ receptor (5-HT₃R) agonists 2-methyl-5-HT, dopamine, and m -chlorophenylbiguanide on 5-HT₃R native to N1E-115 cells and on homopentameric 5-HT₃R expressed in Xenopus oocytes were determined relative to that of 5-HT. Efficacies of 2-methyl-5-HT and dopamine on 5-HT₃R native to differentiated N1E-115 cells are high (54 and 36%) as compared with their efficacies on homopentameric 5-HT₃R-A_L and 5-HT₃R-A_s receptors expressed in oocytes (4–8%). m -Chlorophenylbiguanide does not distinguish between 5-HT₃R in N1E-115 cells and in oocytes. The distinct pharmacological profile of 5-HT₃R native to differentiated N1E-115 cells is conserved when poly(A)⁺ mRNA from these cells is expressed in oocytes. The results indicate that, apart from the known 5-HT₃R subunits, N1E-115 cells express additional proteins involved in 5-HT₃R function.     

A Single Amino Acid Difference Accounts for the Pharmacological Distinctions Between the Rat and Human 5-Hydroxytryptamine1B Receptors

Abstract: Molecular cloning of the rat and human 5-hydroxytryptamine_1B (5-HT_1B) receptors has revealed that the primary amino acid sequence of these two receptors is >90% identical. Despite this high degree of primary sequence homology, these two receptors have significantly different pharmacological properties. A mutant human 5-HT_1B receptor was constructed in which Thr³⁵⁵ was replaced by Asn, the corresponding residue at this position in the rat 5-HT_1B receptor. The pharmacology of the mutant human 5-HT_1B receptor was very similar to that of the rat 5-HT_1B receptor. Specifically, the mutant receptor had much higher affinity for pindolol, [¹²⁵I]-iodocyanopindolol, propranolol, and CP-93,129 than the wild-type receptor. In contrast, the mutant had significantly lower affinity for sumatriptan, N,N -dipropyl-5-carboxamidotryptamine, 5-methoxy- N,N -dimethyltryptamine, methysergide, metergoline, and rauwolscine. These data suggest that a single amino acid difference at position 355 is responsible for the pharmacological differences between the rat and human 5-HT_1B receptors.     

Regulation of Serotonin2A Receptor Expression by an Antisense Oligodeoxynucleotide

Abstract: The regulation of 5-HT_2A receptor expression by an antisense oligodeoxynucleotide, complementary to the coding region of rat 5-HT_2A receptor mRNA, was examined in a cortically derived cell line and in rat brain. Treatment of A₁A₁ variant cells, which express the 5-HT_2A receptor coupled to the stimulation of phosphatidylinositol (PI) hydrolysis, with antisense oligodeoxynucleotide decreased the maximal stimulation of PI hydrolysis by the partial agonist quipazine and the number of 5-HT_2A receptor sites as measured by the binding of 2-[¹²⁵I]-iodolysergic acid diethylamide. Treatment of cells with random, sense, or mismatch oligodeoxynucleotide did not alter the stimulation of PI hydrolysis by quipazine or 5-HT_2A receptor number. Intracerebroventricular infusion of antisense, but not mismatch, oligodeoxynucleotide for 8 days resulted in a significant increase in cortical 5-HT_2A receptor density and an increase in headshake behavior induced by the 5-HT₂ receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane. The density of cortical 5-HT_2A receptors was not altered by administration of antisense oligodeoxynucleotide for 1, 2, or 4 days. We hypothesize that in brain this antisense oligodeoxynucleotide relieved some form of translational suppression, resulting in an increase in 5-HT_2A receptor expression.     

The Third Intracellular Loop of the 5-Hydroxytryptamine2A Receptor Determines Effector Coupling Specificity

Abstract: 5-Hydroxytryptamine (5-HT) receptors contain seven putative transmembrane domains and couple via different guanine nucleotide binding proteins to specific effector enzymes. Studies with other receptors identify the second and third intracellular loops or the C-terminus of the receptor as important for selective effector coupling. However, it is not known which regions of the 5-HT receptor determine effector coupling specificity. To address this question, we constructed a chimeric 5-HT receptor in which the third intracellular (i3) loop is derived from the 5-HT_2A receptor, which is coupled to activation of phospholipase C, and the rest of the sequence is derived from the 5-HT_1B receptor, which is coupled to inhibition of adenylyl cyclase. The chimeric receptor exhibited ligand binding properties similar to those of the 5-HT_1B receptor and distinct from those of the 5-HT_2A receptor. This suggests that the i3 loop is not critical for the unique pharmacology of the 5-HT_1B receptor. In contrast, the chimeric receptor exhibited signaling properties similar to those of the 5-HT_2A receptor and distinct from those of the 5-HT_1B receptor. This indicates that the i3 loop determines the effector coupling specificity of the 5-HT_2A receptor.     

Lack of CB1 receptor activity impairs serotonergic negative feedback

Serotonergic and endocannabinoid systems are important substrates for the control of emotional behaviour and growing evidence show an involvement in the pathophysiology of mood disorders. In the present study, the absence of the activity of the CB₁ cannabinoid receptor impaired serotonergic negative feedback in mice. Thus, in vivo microdialysis experiments revealed increased basal 5-HT extracellular levels and attenuated fluoxetine-induced increase of 5-HT extracellular levels in the prefrontal cortex of CB₁ knockout compared with wild-type mice. These observations could be related to the significant reduction in the 5-HT transporter binding site density detected in frontal cortex and hippocampus of CB₁ knockout mice. The lack of CB₁ receptor also altered some 5-HT receptors related to the 5-HT feedback. Extracellular recordings in the dorsal raphe nucleus (DRN) revealed that the genetic and pharmacological blockade of CB₁ receptor induced a 5-HT_1A autoreceptor functional desensitization. In situ hybridization studies showed a reduction in the expression of the 5-HT_2C receptor within several brain areas related to the control of the emotional responses, such as the DRN, the nucleus accumbens and the paraventricular nucleus of the hypothalamus, whereas an over-expression was observed in the CA3 area of the ventral hippocampus. These results reveal that the lack of CB₁ receptor induces a facilitation of the activity of serotonergic neurons in the DRN by altering different components of the 5-HT feedback as well as an increase in 5-HT extracellular levels in the prefrontal cortex in mice.     

Guinea pig hippocampal 5-HT1E receptors: a tool for selective drug development

Recent studies have indicated that the serotonin [5-hydroxytryptamine (5-HT)] 1E receptor, originally discovered in human brain tissue, is not expressed in rat or mouse brain. Thus, there have been few reports on 5-HT_1E receptor drug development. However, expression of 5-HT_1E receptor mRNA has been shown in guinea pig brain. To establish this species as an animal model for 5-HT_1E drug development, we identified brain regions that exhibit 5-carboxyamidotryptamine, ritanserin, and LY344864 – insensitive [³H]5-HT binding (characteristic of the 5-HT_1E receptor). In hippocampal homogenates, where 5-HT_1E receptor density was sufficiently high for radioligand binding analysis, 100 nM 5-carboxyamidotryptamine, 30 nM ritanserin, and 100 nM LY344864 were used to mask [³H]5-HT binding at non-5-HT_1E receptors. The K _d of [³H]5-HT was 5.7 ± 0.7 nM and is indistinguishable from the cloned receptor K _d of 6.5 ± 0.6 nM. The affinities of 16 drugs for the cloned and hippocampal-expressed guinea pig 5-HT_1E receptors are essentially identical ( R ² = 0.97). These findings indicate that using these conditions autoradiographical distribution and signal transduction studies of the 5-HT_1E receptor in guinea pig brain are feasible. Using the guinea pig as an animal model should provide important insights into possible functions of this receptor and the therapeutic potential of selective human 5-HT_1E drugs.     

Flesinoxan Dose-Dependently Reduces Extracellular 5-Hydroxytryptamine (5-HT) in Rat Median Raphe and Dorsal Hippocampus Through Activation of 5-HT1A Receptors

Abstract: The effects of systemic administration of the serotonin (5-hydroxytryptamine) 5-HT_1A receptor agonists flesinoxan and 8-hydroxy-2-(di- n -propylamino)tetralin on extracellular 5-HT were measured using microdialysis probes in both median raphe nucleus and dorsal hippocampus. Both 5-HT_1A agonists dose-dependently decreased dialysate 5-HT levels from both brain regions. The effects of flesinoxan in the median raphe (0.3 mg/kg) and dorsal hippocampus (1.0 mg/kg) could be blocked by the 5-HT_1A receptor antagonist N -[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]- N -(2-pyridyl)cyclohexane carboxamide trihydrochloride (WAY 100,635) at a dose of 0.05 mg/kg s.c. The antagonist itself had no effect at this dosage. Local perfusion of flesinoxan for 30 min through the dialysis probe into the median raphe region at concentrations of 20, 100, and 1,000 n M resulted in a significant decrease in dialysate 5-HT content from both regions. The effect of 100 n M flesinoxan could be blocked by coperfusion of 1,000 n M WAY 100,635. The data indicate that flesinoxan is a potent 5-HT_1A receptor agonist and also support the notion that somatodendritic 5-HT_1A autoreceptors regulate both terminal and somatodendritic 5-HT release.     

Differential Membrane Targeting and Pharmacological Characterization of Chimeras of Rat Serotonin 5-HT1A and 5-HT1B Receptors Expressed in Epithelial LLC-PK1 Cells

Abstract: The serotonin 5-HT_1A and 5-HT_1B receptors are two structurally related but pharmacologically distinguishable 5-HT receptor types. In brain, the 5-HT_1A receptor is localized on the soma and dendrites of neurons, whereas the 5-HT_1B receptor is targeted to the axon terminals. We previously showed that these two receptors are targeted in different membrane compartments when stably expressed in the epithelial LLC-PK1 cell line. Further investigations on the mechanisms responsible for their differential targeting were done by constructing chimeras of 5-HT_1A and 5-HT_1B receptors still able to bind specifically [³H]lysergic acid diethylamide and selective agonists and antagonists. Their cellular localization examined by confocal microscopy suggests that the third intracellular domain of the 5-HT_1B receptor was responsible for its Golgi-like localization in transfected LLC-PK1 cells. In contrast, the third intracellular domain of the 5-HT_1A receptor apparently allowed the sorting of the chimeras to the plasma membrane. Further inclusion of the C-terminal domain of the 5-HT_1A receptor in their sequence led to a basolateral localization, whereas that of the 5-HT_1B receptor allowed an apical targeting, suggesting the existence of a targeting signal in this portion of the receptor(s).