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Biomedical subjects

B L Roth

Publications and source records attributed to B L Roth.

At least 55 records · Page 3Linked to original sources
PubMed1997

Identification of conserved aromatic residues essential for agonist binding and second messenger production at 5-hydroxytryptamine2A receptors.

Several models of agonist binding to G protein-coupled 5-hydroxytryptamine [5-HT] (serotonin) receptors have highlighted the potential importance of highly conserved aromatic residues for ligand binding and agonist efficacy. In this study, we tested these models by constructing and characterizing a number of point mutations of conserved and nonconserved aromatic residues using the 5-HT2A receptor as a model system. Mutations of three highly conserved tryptophans (W200A, W336A, and W367A) proposed to reside near the binding pocket markedly reduced agonist affinity and efficacy at 5-HT2A receptors. Mutations of two other highly conserved aromatic residues postulated to be near the agonist binding site (F340L and Y370A) also had dramatic effects on agonist binding and efficacy. Point mutations of neighboring conserved phenylalanines (F339L and F365L) had minimal effects on agonist binding, although the F365L mutation diminished agonist efficacy. Finally, mutations of two nonconserved aromatic residues (F125L and F383A) not predicted to be near the binding pocket had no effects on agonist binding, potency, or efficacy. Our results are best explained by models that suggest that helices III, V, VI, and VII can form a unit of interacting helices in which highly conserved aromatic residues are oriented toward the center of the helical aggregate to form an aromatic pocket. In addition, our novel results identify a series of aromatic residues essential for agonist-induced second messenger production. These results demonstrate that highly conserved aromatic residues residing in neighboring helices provide the optimum environment for both agonist binding and activation of 5-HT2A receptors.

DOM 2,5-Dimethoxy-4-Methylamphetamine↗
PubMed1997

Bacterial viability and antibiotic susceptibility testing with SYTOX green nucleic acid stain.

A fluorescent nucleic acid stain that does not penetrate living cells was used to assess the integrity of the plasma membranes of bacteria. SYTOX Green nucleic acid stain is an unsymmetrical cyanine dye with three positive charges that is completely excluded from live eukaryotic and prokaryotic cells. Binding of SYTOX Green stain to nucleic acids resulted in a > 500-fold enhancement in fluorescence emission (absorption and emission maxima at 502 and 523 nm, respectively), rendering bacteria with compromised plasma membranes brightly green fluorescent. SYTOX Green stain is readily excited by the 488-nm line of the argon ion laser. The fluorescence signal from membrane-compromised bacteria labeled with SYTOX Green stain was typically > 10-fold brighter than that from intact organisms. Bacterial suspensions labeled with SYTOX Green stain emitted green fluorescence in proportion to the fraction of permeabilized cells in the population, which was quantified by microscopy, fluorometry, or flow cytometry. Flow cytometric and fluorometric approaches were used to quantify the effect of beta-lactam antibiotics on the cell membrane integrity of Escherichia coli. Detection and discrimination of live and permeabilized cells labeled with SYTOX Green stain by flow cytometry were markedly improved over those by propidium iodide-based tests. These studies showed that bacterial labeling with SYTOX Green stain is an effective alternative to conventional methods for measuring bacterial viability and antibiotic susceptibility.

Ampicillin Resistance↗
PubMed1997

Development of the FUN-1 family of fluorescent probes for vacuole labeling and viability testing of yeasts.

A new family of fluorescent probes has been developed for assessing the viability and metabolic activity of yeasts. This class of halogenated unsymmetric cyanine dyes is exemplified by the FUN-1 [2-chloro-4-(2,3-dihydro-3-methyl-(benzo-1,3-thiazol-2-yl)- methylidene)-1-phenylquinolinium iodide] stain, a membrane-permeant nucleic acid-binding dye that has been found to give rise to cylindrical intravacuolar structures (CIVS) in Saccharomyces cerevisiae. Biochemical processing of the dye by active yeasts yielded CIVS that were markedly red shifted in fluorescence emission and therefore spectrally distinct from the nucleic acid-bound form of the dye. The formation of CIVS occurred under both aerobic and anaerobic conditions and was highly temperature dependent. Treatment of yeasts with the nonmetabolizable glucose analog 2-deoxy-D-glucose reduced cellular ATP levels approximately 6-fold and completely inhibited CIVS formation. Under aerobic conditions, the formation of CIVS was abrogated by the cytochrome oxidase inhibitors azide and cyanide; however, the H+ transport uncoupler carbonyl cyanide m-chlorophenylhydrazone inhibited CIVS formation under both aerobic and anaerobic conditions. Depletion of cellular thiols, including glutathione, with millimolar concentrations of N-ethylmaleimide, iodoacetamide, or allyl alcohol completely inhibited CIVS production. Marked reduction in the formation of CIVS by ethacrynic acid and sulfobromophthalein, inhibitors of glutathione S-transferase, suggested that dye processing can involve enzyme-mediated formation of glutathione conjugates. The conversion of FUN-1 by S. cerevisiae was studied quantitatively by using several techniques, including fluorometry, flow cytometry, and wide-field and confocal laser scanning fluorescence microscopy.

1-Propanol↗
PubMed1997

High-affinity agonist binding is not sufficient for agonist efficacy at 5-hydroxytryptamine2A receptors: evidence in favor of a modified ternary complex model.

In this study, the relationship between high-affinity agonist binding and second messenger production was examined at native and mutant 5-hydroxytryptamine2A receptors. At native 5-hydroxytryptamine2A receptors all agonists, with the exception of quipazine, discriminated between high- and low-affinity states of the receptor, as determined by analysis of competition binding assays. There was no correlation between the ability of selected agonists to label the high-affinity agonist state and to augment phosphoinositide hydrolysis. Quipazine, which did not discriminate between the affinity states of the receptor, behaved as a full agonist. Similar results were obtained when a point mutation (F340L) of a highly conserved phenylalanine located in transmembrane domain VI was examined. With the F340L mutant, most of the agonists tested labeled significantly fewer high-affinity sites, compared with the native receptor. There was no significant relationship between high-affinity agonist binding and second messenger production. Bufotenine and 4-iodo-3,5-dimethoxyphenylisopropylamine labeled similar percentages of high-affinity agonist binding sites (22% vs. 26%), but 4-iodo-3,5-dimethoxyphenylisopropylamine behaved as a full agonist, whereas bufotenine was devoid of detectable agonist activity. The inability of selected agonists to activate phosphoinositide hydrolysis was not due solely to lower agonist affinity for the mutant receptor, because the binding affinity of quipazine was unchanged by the F340L mutation but quipazine had no detectable agonist activity at the mutant receptor. Our results demonstrate that the ability of an agonist to promote the high-affinity state of the 5-hydroxytryptamine2A receptor is not correlated with its ability to augment second messenger production. These results are consistent with recent models of G protein-receptor functioning (e.g., modified ternary complex model) that predict that additional transition states of the receptor-ligand complex are essential for agonist efficacy.

DOM 2,5-Dimethoxy-4-Methylamphetamine↗
PubMed1996

Interactions of selective serotonin reuptake inhibitors with the serotonin 5-HT2c receptor.

Interactions of the selective serotonin reuptake inhibitors (SSRIs) citalopram, fluoxetine and its main metabolite norfluoxetine, and the tricyclic antidepressant (TCA) imipramine with the rat serotonin 5-HT2C receptor in a clonal cell line and in the rat choroid plexus were investigated by radioligand binding and phosphoinositide (PI) hydrolysis assays. For comparison, the affinities of a variety of other antidepressants of different chemical classes for the cloned rat 5-HT2C and 5-HT2A receptors were also determined by radioligand binding assays. Fluoxetine displayed relatively high affinity for the 5-HT2C receptor in the choroid plexus, with a Ki value for inhibition of [3H]mesulergine binding of 55.4 nM. The Ki values for imipramine, norfluoxetine and citalopram were 136 nM, 203 nM, and 298 nM, respectively. Similar rank order of potency was detected in PI hydrolysis assays, which showed that these drugs are antagonists at the 5-HT2C receptor without exhibiting inverse agonist activity. [3H]Ketanserin (5-HT2A) binding assays revealed that the SSRIs fluoxetine, norfluoxetine and citalopram show 10- to 23-fold selectivity for the 5-HT2C receptor in vitro, whereas the TCA imipramine does not. Many other TCAs also had high to intermediate affinity for both 5-HT2A and 5-HT2C receptors. The present data provide evidence that fluoxetine, norfluoxetine and citalopram, along with many other antidepressant compounds, interact directly with the 5-HT2C receptor.

Animals↗
PubMed1996

Cloning, characterization, and chromosomal localization of a human 5-HT6 serotonin receptor.

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-HT1D alpha receptor, suggesting that these may be closely linked. Comparison of genomic and cDNA clones for the human 5-HT6 receptor also reveals an Rsal restriction fragment length polymorphism within the coding region.

Adenylyl Cyclases↗
PubMed1996

Rapid agonist-induced internalization of the 5-hydroxytryptamine2A receptor occurs via the endosome pathway in vitro.

The mechanism by which agonists induce 5-hydroxytryptamine2A (5-HT2A) receptor internalization was investigated in a clonal cell line stably transfected with the 5-HT2A receptor cDNA. Confocal laser microscopy of immunolabeled 5-HT2A receptors in control (untreated) cells demonstrated that most of the immunoreactivity was associated with the cell surface. After quipazine administration, a significant increase in intracellular immunofluorescence was measured. Time course studies demonstrated rapid agonist-dependent internalization of 5-HT2A receptors, with significant internalization occurring as early as 5 min after agonist administration at 37 degrees. In GF-62 cells, agonist-induced internalization was blocked by preincubation with the 5-HT2A receptor antagonist ketanserin. Internalization was also temperature sensitive because agonist-induced internalization did not occur at 4 degrees. Dual-label experiments disclosed that 5-HT2A and transferrin receptors were internalized via the same endocytotic vesicles. These results suggest that 5-HT2A receptors and transferrin receptors are internalized via the endosomal pathway in GF-62 cells. Although 5-HT2A receptors were internalized, down-regulation, or loss of radioligand binding sites, did not occur. Our results demonstrate that agonists rapidly induce 5-HT2A receptor internalization via the endosomal pathway and that internalization can be dissociated from down-regulation.

3T3 Cells↗
PubMed1995

D4 dopamine receptor binding affinity does not distinguish between typical and atypical antipsychotic drugs.

The affinities of 13 atypical and 12 typical antipsychotic drugs for the cloned rat D4 dopamine receptor and the D4/D2 ratios were examined. Of the atypical antipsychotic drugs tested, only clozapine, risperidone, olanzapine, zotepine and tiospirone had affinities less than 20 nM. In fact, many atypical antipsychotic drugs had relatively low affinities for the cloned rat D4 receptor, with Ki values greater than 100 nM (Seroquel, fluperlapine, tenilapine, FG5803 and melperone). Additionally, several typical antipsychotic drugs had high affinities for the cloned rat D4 receptor, with Kis less than 20 nM (loxapine, chlorpromazine, fluphenazine, mesoridazine, thioridazine and trifluoroperazine). The ratios of D2/D4 affinities did not differentiate between these two types of antipsychotic drugs. Thus, D4 dopamine receptor affinity, used as a single measure, does not distinguish between the group of typical and atypical antipsychotic drugs analyzed.

Animals↗
PubMed1995

Differential ergoline and ergopeptine binding to 5-hydroxytryptamine2A receptors: ergolines require an aromatic residue at position 340 for high affinity binding.

In this paper we show that a highly conserved aromatic residue, phenylalanine at the 340-position, is essential for ergoline binding to 5-hydroxytryptamine2A receptors. We hypothesized that F340 was essential for a specific aromatic-aromatic interaction (e.g., pi-pi or hydrophobic) between the phenyl moiety of F340 and the aromatic ring of the ergoline nucleus. To test this hypothesis, eight point mutations of adjacent (F340 and F339) and nonadjacent (F125) phenylanaines were made, using conservative (phenylalanine to tyrosine) and nonconservative (phenylalanine to leucine, alanine, or serine) substitutions. The binding affinities of all of the tested simple ergolines were greatly reduced by specific mutations of F340 in which aromatic-aromatic interactions (e.g., F340A and F340L) were abolished, but they were unaffected when the replacement residue was aromatic (e.g., F340Y). In contrast, the binding affinities of four ergopeptines (bromocryptine, ergocryptine, ergocornine, and ergotamine) were relatively unaffected by the F340L substitution. Neither ergoline nor ergopeptine affinities were consistently altered by F339 mutations. These results support the notion that aromatic-aromatic interactions (either pi-pi of hydrophobic) with F340 are essential for the binding of simple ergolines but not ergopeptines to 5-hydroxytryptamine2A receptors. Our findings support models of ergoline and ergopeptine binding to serotonin receptors that suggest that the nature of the substituent at the 8-position of the ergoline nucleus may give rise to different modes of binding for the two classes of agents, particularly with respect to the phenyl ring of F340.

Animals↗
PubMed1995

5-Hydroxytryptamine2A (5-HT2A) receptor desensitization can occur without down-regulation.

The connection between agonist-induced desensitization and down-regulation of 5-hydroxytryptamine2A (5-HT2A) receptors was examined in a clonal cell line that stably expresses the 5-HT2A receptor. Brief (2-hr) and prolonged (24-hr) exposure to the agonist quipazine or the agonist 4-iodo-(2,5-dimethoxy)- phenylisopropylamine (DOI) diminished 5-HT2A receptor-mediated phosphoinositide hydrolysis; no change in 5-HT2A receptor number or affinity was measured after 24 hr of exposure to DOI or quipazine. Immunohistochemical studies demonstrated that a 24-hr exposure to DOI did not alter surface 5-HT2A receptor immunoreactivity. Western blot analysis with G alpha q- and G alpha 11-selective antibodies indicate that a 24-hr agonist exposure did not alter the levels of phospholipase C-dependent G proteins. These results suggest that desensitization after prolonged DOI exposure can occur via a process independent of the levels of phospholipase C-coupled G proteins. Studies with a mutant 5-HT2A receptor (F340L) indicated that binding per se is not sufficient for desensitization. Down-regulation of the protein kinase C isozymes alpha and epsilon by overnight exposure to phorbol-12,13-dibutyrate attenuated the intermediate phase (i.e., after 2-6 hr of agonist exposure) of DOI- and quipazine-induced desensitization. These results indicate that the intermediate phase of DOI-induced desensitization is mediated by the alpha- and/or epsilon-protein kinase C isozymes but that neither is involved in the later phase (i.e., after 24 hr of agonist exposure) of desensitization.(ABSTRACT TRUNCATED AT 250 WORDS)

3T3 Cells↗
PubMed1994

1-(2,5-Dimethoxy-4-(trifluoromethyl)phenyl)-2-aminopropane: a potent serotonin 5-HT2A/2C agonist.

A method was found to synthesize 1-(2,5-dimethoxy-4-(trifluoromethyl) phenyl)-2-aminopropane, 5, and its des-alpha-methyl congener 2-(2,5-dimethoxy-4-(trifluoromethyl)phenyl)aminoethane, 6, the trifluoromethyl analogs of substituted hallucinogenic phenethylamine derivatives such as 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (3, DOI) that are potent 5-HT2A/2C agonists. In our hands, 5 and 6 have proven to have affinity for [3H]ketanserin or [125I]-3-labeled 5-HT2A/2C sites in rat cortex comparable to or higher than the analogous bromo or iodo analogs. Similarly, 5 and 6 had potency comparable to or slightly greater than that of their bromo or iodo congeners in the two-lever drug discrimination assay in rats trained to discriminate saline from LSD tartrate. The agonist properties of 5 and 6 were evaluated by measuring the accumulation of [3H]inositol monophosphate in cultured cells selectively expressing either 5-HT2A or 5-HT2C receptors. In comparison to serotonin (5-HT), compounds 3 (DOI), 5, and 6 were equally efficacious and full agonists at the 5-HT2C receptor. Similarly, 3 and 5 produced equivalent responses at the 5-HT2A receptor as compared to 5-HT. In contrast, 6, the alpha-desmethyl analog of 5, was only half as potent at stimulating inositol monophosphate accumulation at the 5-HT2A receptor. In conclusion, the title compound 5 and its alpha-desmethyl congener 6 appear to be the most potent of the so-called hallucinogenic amphetamine 5-HT agonists reported to date. Further, the reduced efficacy of 6 at the 5-HT2A receptor may offer at least a partial explanation for the observed higher in vivo potencies of alpha-methyl-substituted compounds in this series.

3T3 Cells↗
PubMed1994

Effect of the R(-) and S(+) isomers of MDA and MDMA on phosphatidyl inositol turnover in cultured cells expressing 5-HT2A or 5-HT2C receptors.

The effect of the R(-) and S(+) isomers of 3,4-methylenedioxyamphetamine (MDA) and its N-methyl analog 3,4-methylenedioxymethamphetamine (MDMA) on [3H]inositol monophosphate accumulation was studied in cells expressing either 5-HT2A or 5-HT2C receptors. The isomers of MDA produced a concentration dependent increase in phosphatidyl inositol (PI) hydrolysis at the 5-HT2A receptors, with the R(-) isomer of MDA being more potent than the S(+) at the 5-HT2A receptor. The R(-) and S(+) isomers of MDMA were significantly less efficacious at the 5-HT2A receptor as compared to MDA; S(+)MDMA had no effect. At the 5-HT2C receptor, both R(-) and S(+)MDA were equipotent at stimulating PI hydrolysis, with the S(+) isomer of MDMA being more efficacious at the 5-HT2C receptor compared with the R(-) isomer. In all cases at both the 5-HT2A and 5-HT2C receptors, the affinities of the isomers of MDMA and MDA were at least 2-3 orders of magnitude less than 5-HT. Despite the weak effect of these compounds at the 5-HT2A and 5-HT2C receptors, these substituted amphetamines do possess intrinsic activity which may contribute to their neurotoxic effects when administered at high doses.

3,4-Methylenedioxyamphetamine↗
PubMed1994

Multiple serotonin receptors: clinical and experimental aspects.

Serotonin (5-hydroxytryptamine; 5-HT) has been implicated in a large number of psychophysiologic processes including the regulation of sleep, appetite, mood, aggression, perception, memory, and anxiety. To mediate this large array of physiologic processes, at least 14 separate 5-HT receptors have evolved, which are divided into seven main families. Not surprisingly, alterations of 5-HT receptor activity have been shown to occur in many psychiatric diseases including anxiety, depression, eating disorders, schizophrenia, personality disorders, and many drug-induced psychotic states. Additionally, a number of effective psychopharmacologic agents for diseases as diverse as schizophrenia and anxiety have been developed which either specifically alter brain levels of serotonin or bind to 5-HT receptor subtypes. This review article summarizes recent advances in the burgeoning field of serotonin receptor pharmacology and integrates this information into a coherent perspective on the importance of serotonergic agents for clinical psychiatry.

Animals↗
PubMed1994

Binding of typical and atypical antipsychotic agents to 5-hydroxytryptamine-6 and 5-hydroxytryptamine-7 receptors.

The authors examined the affinities of 36 typical and atypical antipsychotic agents for the cloned rat 5-hydroxytryptamine-6 (5-HT6) and rat 5-hydroxytryptamine-7 (5-HT7) receptors in transiently expressed COS-7 cells (5-HT7) or stably transfected HEK-293 cells (5-HT6 receptors). Clozapine and several related atypical antipsychotic agents (rilapine, olanzepine, tiospirone, fluperlapine, clorotepine and zotepine) had high affinities for the newly discovered 5-HT6 receptor (Kis < 20 nM). The 5-HT7 receptor bound clozapine, rilapine, fluperlapine, clorotepine, zotepine and risperidone but not tiospirone and olanzepine, with affinities less than 15 nM. In addition, several typical antipsychotic agents (chloroprothixene, chlorpromazine, clothiapine and fluphenazine) had high affinities for both the 5-HT6 and 5-HT7 receptors. Pimozide, a diphenylbutylpiperidine, had the highest affinity of all the typical antipsychotic agents tested for the 5-HT7 receptor (Ki = 0.5 nM). Three putative atypical antipsychotic agents melperone, amperozide and MDL 100907 did not bind with high affinities to either the 5-HT6 or 5-HT7 receptors (Kis > 50 nM). Several dopamine-selective antipsychotic agents (raclopride, rimcazole and penfluridol) had essentially no affinity for either the 5-HT6 or 5-HT7 receptors (Ki values > 5000 nM).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Production and characterization of a specific 5-HT2 receptor antibody.

A synthetic peptide was used to generate antibodies against the rat serotonin-2 (5-HT2) receptor. The peptide corresponds to a unique sequence from the N-terminal extracellular portion of the receptor protein (antibody = Ab 5HT2-N). This peptide was chosen based on its theoretical antigenic index and for specificity to the 5-HT2 receptor. In dot blot analysis, antisera detected 2 ng-2 micrograms of synthetic peptide at dilutions of 1/200-1/20,000. COS-7 cells transiently transfected with a eukaryotic expression vector containing the 5-HT2 cDNA displayed intense immunoreactivity with crude and affinity-purified Ab 5HT2-N. In contrast, no immunoreactivity was seen in control experiments when: (1) non-transfected or vector transfected COS-7 cells were used; (2) pre-immune sera was substituted for primary antisera; (3) primary antisera was omitted; or (4) antiserum was pre-adsorbed to 10 microM synthetic peptide. Immunohistochemical analysis of sections of perfused rat brain revealed intense immunolabelling of a subset of neurons in regions of the ventral forebrain, dorsal hippocampus, striatum, cerebral cortex, and laterodorsal tegmental nucleus (LDT). An especially dense band of small cells was seen in layer 2 of pyriform cortex. There was a very high concentration of labelled cells in the laterodorsal tegmental nucleus. In situ hybridization histochemistry with a 5-HT2 antisense cRNA riboprobe showed a pattern of hybridization in forebrain similar to the pattern of immunolabelling with Ab 5HT2-N. Western blot analysis of proteins extracted from the LDT revealed a single protein species reacting with the antibody. This reactivity is not present in the pre-immune sera and is blocked by the synthetic antigen.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1993

A single point mutation (Phe340-->Leu340) of a conserved phenylalanine abolishes 4-[125I]iodo-(2,5-dimethoxy)phenylisopropylamine and [3H]mesulergine but not [3H]ketanserin binding to 5-hydroxytryptamine2 receptors.

The molecular processes by which agonists and antagonists bind to serotonin2 [5-hydroxytryptamine (5-HT2)] receptors are currently unknown. Three molecular models have proposed that conserved aromatic residues help to anchor the phenyl ring of 5-HT via stacking or pi-pi-type interactions with the 5-HT2 receptor. To test these models we made single point mutations (Phe339-->Leu339 and Phe340-->Leu340) of two aromatic residues that are conserved among all guanine nucleotide-binding protein-coupled 5-HT receptors and a single point mutation (Phe125-->Leu125) that exchanges a 5-HT2 for a 5-HT1c sequence. [3H]Mesulergine binding was abolished by Phe340-->Leu340 and unchanged with the Phe339-->Leu339 and Phe125-->Leu125 mutations, whereas [3H]ketanserin binding affinity was diminished by the Phe339-->Leu339 mutation and unchanged by Phe340-->Leu340 and Phe125-->Leu125. We also found that the affinities of three ergot derivatives (mesulergine, methysergide, and lisuride) were decreased by 88-1079-fold with only the Phe340-->Leu340 mutation. We also discovered that 4-[125I]iodo-2,5-(dimethoxy)phenylisopropylamine (DOI) binding was abolished in COS-7 cells expressing 5-HT2 (Phe340-->Leu340) receptors but maintained in cells expressing the Phe339-->Leu339 and Phe125-->Leu125 mutations. Additionally, the Ki values for several agonists and partial agonists (5-HT, DOI, m-chlorophenylpiperazine, trifluoromethylphenylpiperazine, bufotenine, and MK-212) were greatly diminished (26-14,000-fold decrease) only with the Phe340-->Leu340 receptor mutation. Finally, the Phe340-->Leu340 mutant receptors displayed an attenuated or abolished ability to augment phosphoinositide hydrolysis in COS-7 cells with four separate agonists (5-HT, MK-212, bufotenine, and quipazine). Taken together, these results are consistent with the idea that agonists and certain ergot derivatives anchor to 5-HT2 receptors, in part, via specific interactions with aromatic residue Phe340 located in transmembrane region VI.

Amino Acid Sequence↗
PubMed1992

Binding of typical and atypical antipsychotic agents to transiently expressed 5-HT1C receptors.

We determined the affinities of clozapine and 21 other typical and atypical antipsychotic agents for the cloned 5-hydroxytryptamine-1C (5-HT1C) receptor. For these studies, 5-HT1C receptors were transiently expressed in COS-7 cells using the vector pSVK3-5HT1C. We discovered that clozapine and several other putative typical and atypical antipsychotic agents (loxapine greater than tiosperone greater than SCH23390 greater than fluperlapine greater than rilapine greater than chlorpromazine) had relatively high affinities (7-30 nM) for the cloned 5-HT1C receptor. Other antipsychotic agents (risperidone greater than tenilapine greater than mesoridazine greater than thioridazine greater than cis-fluphenthixol) had intermediate affinities (30-100 nM), whereas many other antipsychotics (fluphenazine greater than spiperone greater than amperozide greater than melperone greater than thiothixene greater than haloperidol, metoclopramide, pimozide, domperidone, sulpiride) had low affinities (greater than 500 nM) for the cloned 5-HT1C receptor. The results indicate that although several putative atypical antipsychotic agents have high affinities for the cloned rat 5-HT1C receptor, the spectrum of drug binding does not correlate with the atypical nature of these compounds.

Antipsychotic Agents↗