Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

PCR was used to isolate identical partial cDNA clones encoding a serotonin 5-HT3 receptor subunit from rat nodose and superior cervical ganglia. The amino acid sequence predicted from these clones, extending from the putative transmembrane domain I to the stop codon, demonstrated a 93% homology with the 5-HT3 receptor A (R-A) subunit cloned from NCB 20 hybridoma mouse neuroblastoma/Chinese hamster embryonic brain cells. Comparison of the sequences of the rat gene and cDNA encoding this subunit revealed a five amino acid deletion, GSLLP, located within the putative second intracellular loop of the receptor subunit. This deletion was shown to occur at an intron/exon junction. Therefore, alternative splicing was probably responsible for the presence of short (5-HT3 R-As) and long (5-HT3 R-AL) forms of 5-HT3 R-A mRNA in these ganglia. PCR experiments, with specific primers located upstream and downstream of the GSLLP deletion, were used to detect reverse transcribed 5-HT3 R-A mRNAs. A short fragment (92 bp), corresponding to the deleted form, and a long fragment (107 bp), corresponding to the nondeleted form, were amplified from various regions of the CNS and peripheral ganglia of the rat, as well as from NG108-15 hybridoma cells. In the adult rat, the ratio of the two forms varied very little from one tissue to another, the long form corresponding to only approximately 10% of the total 5-HT3 R-A mRNA. Study of their respective distributions during ontogeny demonstrated a differential expression of the short and long forms in some tissues during late embryonic development, at embryonic day 17 (E17) or E20.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Developmental changes in the differential expression of two serotonin 5-HT3 receptor splice variants in the rat. 761

1. The pharmacological and biophysical properties of a recombinant 5-HT3 receptor have been studied by use of patch-clamp techniques applied to HEK 293 cells stably transfected with the murine 5-HT3 R-A cDNA. 2. At a holding potential of -60 mV, 77% of cells investigated responded to ionophoretically applied 5-HT with an inward current. Such currents were unaffected by methysergide (1 microM), or ketanserin (1 microM), but were antagonized in a concentration-dependent and reversible manner by the selective 5-HT3 receptor antagonist, ondansetron (IC50 = 440 pM) and the non-selective antagonists (+)-tubocurarine (IC50 = 1.8 nM) and metoclopramide (IC50 50 nM). 3. The 5-HT-induced current reversed in sign (E5-HT) at approximately -2mV and exhibited inward rectification. The influence of extra- and intracellular ion substitutions upon E5-HT indicates the 5-HT-evoked current to be mainly mediated by a mixed monovalent cation conductance. 4. Calcium and magnesium (0.1-10 nM) produced a concentration-dependent, voltage-independent, inhibition of the 5-HT-induced response. Zinc (0.3-300 microM) exerted a biphasic effect with low concentrations enhancing, and high concentrations depressing, the 5-HT-evoked current. 5. Fluctuation analysis of inward currents evoked by a low (1 microM) concentration of 5-HT suggests the current to be mediated by the opening of channels with a conductance of 420 fS. 6. The pharmacological and biophysical properties of the 5-HT3 R-A are similar to those previously described for 5-HT3 receptors native to murine neuroblastoma cell lines, with the exception that the function of the recombinant receptor was enhanced by low concentrations of zinc. This observation suggests that the properties of the native receptor are not completely represented by the 5-HT3 R-A subunit alone.
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PMID:An electrophysiological investigation of the properties of a murine recombinant 5-HT3 receptor stably expressed in HEK 293 cells. 762 Jul 11

In the presence of substance P (SP; 10 microM), serotonin (5-HT; 1 microM) triggered a cation permeability in cells of the hybridoma (mouse neuroblastoma x rat glioma) clone NG 108-15 that could be assessed by measuring the cell capacity to accumulate [14C]guanidinium for 10-15 min at 37 degrees C. In addition to 5-HT (EC50 0.33 microM), the potent 5-HT3 receptor agonists 2-methyl-serotonin, phenylbiguanide, and m-chlorophenylbiguanide, and quipazine, markedly increased [14C]guanidinium uptake in NG 108-15 cells exposed to 10 microM SP. In contrast, 5-HT3 receptor antagonists prevented the effect of 5-HT. The correlation (r = 0.97) between the potencies of 16 different ligands to mimic or prevent the effects of 5-HT on [14C]guanidinium uptake, on the one hand, and to displace [3H]zacopride specifically bound to 5-HT3 receptors on NG 108-15 cells, on the other hand, clearly demonstrated that [14C]guanidinium uptake was directly controlled by 5-HT3 receptors. Various compounds such as inorganic cations (La3+, Mn2+, Ba2+, Ni2+, and Zn2+), D-tubocurarine, and memantine inhibited [14C]guanidinium uptake in NG 108-15 cells exposed to 5-HT and SP, as expected from their noncompetitive antagonistic properties at 5-HT3 receptors. However, ethanol (100 nM), which has been reported to potentiate the electrophysiological response to 5-HT3 receptor stimulation, prevented the effects of 5-HT plus SP on [14C]guanidinium uptake. The cooperative effect of SP on this 5-HT3-evoked response resulted neither from an interaction of the peptide with the 5-HT3 receptor binding site nor from a possible direct activation of G proteins in NG 108-15 cells. Among SP derivatives, [D-Pro9]SP, a compound inactive at the various neurokinin receptor classes, was the most potent to mimic the stimulatory effect of SP on [14C]guanidinium uptake in NG 108-15 cells exposed to 5-HT. Although the cellular mechanisms involved deserve further investigations, the 5-HT-evoked [14C]guanidinium uptake appears to be a rapid and reliable response for assessing the functional state of 5-HT3 receptors in NG 108-15 cells.
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PMID:Characteristics of [14C]guanidinium accumulation in NG 108-15 cells exposed to serotonin 5-HT3 receptor ligands and substance P. 768 66

The effect of acute exposure to alcohols on ion current mediated by recombinant 5-HT3RA receptors transiently expressed in human embryonic kidney 293 cells was investigated. Cells transfected with 5-HT3RA cDNA expressed receptors with pharmacological and functional properties similar to those of native 5-HT3 receptors. Potentiation of receptor-mediated cation current was observed in the presence of ethanol (10-100 mM), butanol (0.1-20 mM), isopentanol (0.01-25 mM) and trichloroethanol (0.5-25 mM). Potentiation increased in a concentration-dependent manner until saturation was achieved for all alcohols tested. The maximal efficacies of potentiation differed among the alcohols with isopentanol > butanol = trichloroethanol > ethanol. Potentiation by butanol and isopentanol appeared to show acute tolerance such that the percent increase in current amplitude was largest upon the first of a series of alcohol applications and decreased during subsequent applications. The effect of ethanol was variable with potentiation occurring in 74% of cells examined, but not in the remaining cells. These observations indicate that the potentiating action of alcohols is similar in recombinant receptors to that previously observed in neuroblastoma cells and neurons expressing native receptors. These findings indicate that this recombinant system is suitable for studying the molecular basis of alcohol actions on the 5-HT3 receptor.
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PMID:Alcohols potentiate ion current mediated by recombinant 5-HT3RA receptors expressed in a mammalian cell line. 776 Sep 80

We have used single-cell imaging of fura-2-loaded cells to examine the Ca2+ signals evoked by activation of 5-hydroxytryptamine type 3 (5-HT3) receptors in undifferentiated N1E-115 neuroblastoma cells and in human embryonic kidney (HEK) 293 cells transfected with either of the two cloned 5-HT3 receptor subunits. The selective 5-HT3 receptor agonist 1-(m-chlorophenyl)-biguanide (mCPBG) caused a concentration-dependent increase in the cytoplasmic Ca2+ concentration ([Ca2+]i) in N1E-115 cells and in HEK 293 cells transfected with either the 5-HT3 A subunit or the 5-HT3 As subunit. In each case, the [Ca2+]i rise was steeply dependent on the mCPBG concentration (nH = 2-4) and abolished by removal of extracellular Ca2+ or addition of ondansetron. Pretreatment of N1E-115 cells with thapsigargin, caffeine, and ryanodine to deplete intracellular Ca2+ stores had no effect on the mCPBG-evoked Ca2+ signals, indicating that they result entirely from stimulated Ca2+ entry. The steep concentration-effect curves therefore are not a consequence of amplification of Ca2+ influx by Ca(2+)-induced Ca2+ release from intracellular stores and probably reflect cooperative activation of 5-HT3 receptors by mCPBG. Depolarization of transfected HEK 293 cells with medium containing increased K+ concentrations invariably failed to evoke an increase in [Ca2+]i, confirming the absence of voltage-gated Ca2+ channels and indicating that the mCPBG-evoked rise in [Ca2+]i results from Ca2+ permeation of 5-HT3 receptors. However, in N1E-115 cells and transfected HEK 293 cells, both extracellular Na+ and K+ substantially inhibited the Ca2+ influx evoked by activation of 5-HT3 receptors, possibly by inhibition of agonist binding or by competition with Ca2+ for permeation of the channel. We conclude that 5-HT3 receptors are Ca2+ permeant, that the Ca2+ influx is sufficient to generate a significant rise in [Ca2+]i, and that, because the A and As subunits behave similarly, conflicting electrophysiological analyses of Ca2+ currents cannot be explained by differences between these two subunits.
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PMID:Ca2+ permeability of cloned and native 5-hydroxytryptamine type 3 receptors. 780 32

The structure of the mouse 5-HT3 receptor gene, 5-HT3R-A, is most similar to nicotinic acetylcholine receptor (nAChR) genes, in particular to the gene encoding the neuronal nAChR subunit alpha 7. These genes share among other things the location of three adjacent introns, suggesting that 5-HT3R-A and nAChR genes arose from a common precursor gene. The alternative use of two adjacent splice acceptor sites in intron 8 creates, in addition to the original 5-HT3R-A cDNA (5-HT3R-AL), a shorter isoform (5-HT3R-AS) which lacks six codons in the segment that translates into the major intracellular domain. This splice consensus sequence is not found in human genomic DNA. In mouse, we demonstrate by RNAse protection assay that 5-HT3R-AS mRNA is approximately 5 times more abundant than 5-HT3R-AL mRNA in both neuroblastoma cell lines and neuronal tissues. We used the Semliki Forest virus expression system for electrophysiological characterization of 5-HT3R-AS and 5-HT3R-AL in mammalian cells. No differences in electrophysiological characteristics, such as voltage dependence, desensitization kinetics, or unitary conductance were found between homomeric 5-HT3R-AS and 5-HT3R-AL receptors. Their properties are very similar to those of 5-HT3 receptors in mouse neuroblastoma cell lines.
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PMID:Organization of the mouse 5-HT3 receptor gene and functional expression of two splice variants. 785 52

1. The aim of this study was to provide evidence that anpirtoline, which is an agonist at 5-HT1B and 5-HT1D receptors and also displays submicromolar affinity for 5-HT1A recognition sites, in addition, acts as an antagonist at 5-HT3 receptors. 2. In radioligand binding studies on rat brain cortical membranes, anpirtoline inhibited specific binding of [3H]-(S)-zacopride to 5-HT3 receptor recognition sites (pKi: 7.53). 3. In N1E-115 neuroblastoma cells in which [14C]-guanidinium was used as a tool to measure cation influx through the 5-HT3 receptor channel, the 5-HT-induced influx was concentration-dependently inhibited by anpirtoline. In this respect, anpirtoline mimicked other 5-HT3 receptor antagonists; the rank order of potency was ondansetron > anpirtoline > metoclopramide. 4. The concentration-response curve for 5-HT as a stimulator of [14C]-guanidinium influx was shifted to the right by anpirtoline (apparent pA2: 7.78). 5. In urethane-anaesthetized rats, anpirtoline inhibited (at lower potency than zacopride and tropisetron) the 5-HT- or phenylbiguanide-induced bradycardia (Bezold-Jarisch reflex), but did not induce this reflex by itself. 6. Intravenous infusion of cisplatin in the domestic pig caused a consistent emetic response which was antagonized by anpirtoline. 7. It is concluded that anpirtoline, which was previously characterized as a 5-HT1 receptor agonist also proved to be a 5-HT3 receptor antagonist in several experimental models and, hence, exhibits a unique pattern of properties at different 5-HT receptors.
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PMID:5-HT3 receptor antagonism by anpirtoline, a mixed 5-HT1 receptor agonist/5-HT3 receptor antagonist. 788 26

The effects of 24 biguanide and four guanidine derivatives on 5-hydroxytryptamine (5-HT)3 receptors in N1E-115 neuroblastoma cells were examined using radioligand binding and whole-cell voltage-clamp techniques. Displacement of the selective 5-HT3 receptor antagonist [3H]BRL 43694 by phenylbiguanide (PBG) derivatives revealed Ki values ranging from 3.4 x 10(-4) to 4.4 x 10(-10) M. The rank order of potency of agonists was 2,3,5-trichloro-PBG > 2,3-dichloro-PBG = 2,5-dichloro-PBG = 3,5-dichloro-PBG > 3,4-dichloro-PBG = 3-chloro-PBG > 2-chloro-PBG = 4-chloro-PBG = 2-methyl-PBG = 2,4-difluoro-PBG > PBG = 2-trifluoro-5-chloro-PBG > 4-fluoro-PBG = 3-trifluoromethyl-PBG > 4-nitro-PBG = 1,5-bis-4-chloro-PBG = 3,5-ditrifluoromethyl-PBG > 4-ethoxy-PBG >> 4-sulfonic acid-PBG. All of the benzylbiguanides and indanylbiguanide were inactive on [3H]BRL 43694 binding or displaced it only weakly. The four guanidine derivatives were quite inactive. In the PBG series, all antagonist competition curves were steep (pseudo-Hill coefficients ranging from 1.05 to 1.58), monophasic, and best fit with a one-site model. Among PBG derivatives, the chlorinated compounds exhibited a good degree of selectivity for 5-HT3 receptors versus other 5-HT receptor subtypes and other neurotransmitter binding sites. Electrophysiological studies showed that the PBG derivatives tested produced rapid inward currents, at a holding potential of -65 mV, that showed rapid desensitization. The current induced by the 2,3,5-trichloro-PBG derivative was inhibited by the specific 5-HT3 receptor antagonist ICS 205-930 but was unaffected by the 5-HT2 receptor antagonist ketanserin. Analysis of concentration-response curves for the PBG derivatives gave EC50 values ranging from 2.2 x 10(-5) to 2.7 x 10(-8) M and Hill slopes ranging from 1.02 to 2.10. The rank order of potency was similar to that obtained from the binding data, and a good correlation was found between Ki and EC50 values. It is concluded that the triple-chloro substitution yielded a compound that is 30-fold more potent than 3-chloro-PBG and approximately 10-fold more potent than dichloro-PBG derivatives, making 2,3,5-trichloro-PBG the most potent 5-HT3 agonist described thus far.
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PMID:Biguanide derivatives: agonist pharmacology at 5-hydroxytryptamine type 3 receptors in vitro. 796 53

The effects of chlorpromazine and the lidocaine derivative QX222, which block nicotinic acetylcholine receptors, were examined on 5-HT3 receptors in N1E-115 mouse neuroblastoma cells, using whole cell voltage-clamp and radioligand binding. Electrophysiological studies examining the effects of chlorpromazine and QX222 on 5-HT3 agonist-induced responses revealed IC50s of 0.2 and 8.5 microM respectively. The action was not voltage- or use-dependent and there was no blocking action when chlorpromazine was applied from inside the cell. Chlorpromazine and QX222 inhibited the binding of a radiolabelled 5-HT3 receptor antagonist, [3H]GR65630, with IC50s of 0.9 and 29 microM respectively. Scatchard plots revealed a decrease in affinity (Kd) in the presence of chlorpromazine, but no change in the maximum number of binding sites (Bmax). The results suggest differential actions of the compounds at 5-HT3 and acetylcholine receptors.
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PMID:Chlorpromazine and QX222 block 5-HT3 receptors in N1E-115 neuroblastoma cells. 798 88

Stereoselective effects of mianserin and ORG3770 on serotonin 5-HT3 receptors in mouse neuroblastoma N1E-115 cells have been investigated in radioligand binding and in whole-cell voltage clamp experiments. The specific binding of [3H]GR65630 to 5-HT3 recognition sites in N1E-115 cell homogenates is reduced by mianserin and ORG3770 and their enantiomers. The pKi values of the more potent (R)enantiomers of mianserin and ORG3770 are 8.44 and 8.62, respectively. The (R)enantiomers of mianserin and ORG3770 are 15 and 37 times more potent than their respective (S)enantiomers. The racemates are only 1.9 and 3.3 times less potent than the corresponding (R)enantiomers. In voltage clamp experiments the (R)enantiomers block the 5-hydroxytryptamine(5-HT)-induced ion current with pIC50 values of 8.52 for (R)mianserin and 8.26 for the (R)enantiomer of ORG3770. The (R)enantiomers of mianserin and ORG3770 are 24 and 145 times more potent in blocking the 5-HT-induced ion current than their respective (S)enantiomers. The racemates are 6 and 13 times less potent than the corresponding (R)enantiomers. In addition, the block of 5-HT-induced ion current by the (R)enantiomer of ORG3770 is partially reversed by a low concentration of its (S)enantiomer. The results indicate that the two enantiomers block the 5-HT3 receptor-mediated ion current in a mutually dependent manner.
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PMID:Interaction between enantiomers of mianserin and ORG3770 at 5-HT3 receptors in cultured mouse neuroblastoma cells. 798 89


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