Introduction of the 5-HT3B subunit alters the functional properties of 5-HT3 receptors native to neuroblastoma cells.
The identification of a second 5-HT(3) (5-HT(3B)) subunit provides an explanation for 5-HT(3) receptor heterogeneity. We investigated whether introduction of recombinant 5-HT(3B) subunits would alter the functional properties of mouse neuroblastoma 5-HT(3) receptors. RT-PCR analysis revealed that NB41A3 cells contain mRNAs encoding 5-HT(3A) and 5-HT(3B) subunits. 5-HT increased intracellular Ca(2+) concentration ([Ca(2+)](i)) and caused the concentration-dependent activation of inward currents recorded at -60 mV. Both actions of 5-HT were antagonized by ondansetron. The 5-HT concentration-response relationship of NB41A3 cells was indistinguishable from that of the related NG108-15 cell line. The selective 5-HT(3)-receptor agonist mCPBG also elevated [Ca(2+)](i) and activated inward currents. 2-M-5HT was less efficacious than 5-HT as an activator of 5-HT(3) receptors in NB41A3 cells and did not significantly increase [Ca(2+)](i). The 5-HT induced increase in [Ca(2+)](i) did not involve caffeine- or thapsigargin-sensitive intracellular Ca(2+) stores. The introduction of the 5-HT(3B) subunit by transient transfection of NB41A3 cells caused 5-HT to become less potent as an activator of 5-HT(3) receptors and altered the kinetics of 5-HT activated currents so that they resembled currents mediated by 5-HT(3AB) receptors. The 5-HT(3B) subunit also abolished the 5-HT induced [Ca(2+)](i) increase seen in untransfected NB41A3 cells. These data are consistent with the hypothesis that NB41A3 cells predominantly express homomeric 5-HT(3A) receptors that become heteromeric 5-HT(3AB) receptors upon introduction of the recombinant 5-HT(3B) subunit.
Stewart
A
A
Department of Pharmacology, The George Washington University, 2300 Eye Street NW, Washington DC 20037, USA.
Davies
P A
PA
Kirkness
E F
EF
Safa
P
P
Hales
T G
TG
D000818:Animals; D001645:Biguanides / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line / Q000145:classification; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D005453:Fluorescence; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009447:Neuroblastoma / Q000145:classification; D017294:Ondansetron / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D021122:Protein Subunits / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D011985:Receptors, Serotonin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D044406:Receptors, Serotonin, 5-HT3; D011993:Recombinant Fusion Proteins / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012701:Serotonin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D012702:Serotonin Antagonists / Q000494:pharmacology; D017366:Serotonin Receptor Agonists / Q000494:pharmacology; D014162:Transfection
D001645:Biguanides; D008164:Luminescent Proteins; D021122:Protein Subunits; D012333:RNA, Messenger; D011985:Receptors, Serotonin; D044406:Receptors, Serotonin, 5-HT3; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D012702:Serotonin Antagonists; D017366:Serotonin Receptor Agonists; D049452:Green Fluorescent Proteins; D012701:Serotonin; D017294:Ondansetron; C045918:2-methyl-5-HT; C065734:1-(3-chlorophenyl)biguanide; D002118:Calcium