Abstract
The cDNA from a schizophrenic patient heterozygous for a mutation of the 5-HT1A receptor gene was used to clone the variant and wild-type DNA into a eukaryotic expression vector. The mutation was characterized by a base pair substitution (A → G) at the first position of codon 28, leading to an Ile → Val amino acid exchange. COS-7 cells were transfected with the cDNA of either the wild type or the variant 5-HT1A receptor. The potencies of the 5-HT1A receptor agonists 8-hydroxy-2-(di-n-propylamino)tetraline (8-OH-DPAT), 5-HT and roxindole, and of the antagonists methiothepin and spiperone in inhibiting specific binding of [3H]8-OH-DPAT of the mutant and wild-type 5-HT1A receptor were determined. All five 5-HT1A receptor ligands concentration-dependently inhibited specific [3H]8-OH-DPAT binding to both the wild-type and the variant 5-HT1A receptor. The rank order of potency of the ligands in inhibiting [3H]8-OH-DPAT binding was identical at both receptors and was roxindole > 8-OH-DPAT > 5-HT > methiothepin > spiperone. This rank order is characteristic for 5-HT1A receptors. The negative logarithms of the concentrations required for 50% inhibition (pIC50 values) of the ligands at the mutant 5-HT receptor correlated highly significantly with those at the wild-type receptor (r = 0.995). It is concluded that the pharmacological profile of the mutant 5-HT1A receptor does not differ from that of the wild-type 5-HT1A receptor.
References
Adham N, Tamm JA, Salon JA, Vaysse PJ-J, Weinshank RL, Brancheck TA (1994) A single point mutation increases the affinity of serotonin 5-HT1Dα, 5-HT1Dβ, 5-HT1E and 5-HT1F receptors for β-adrenergic antagonists. Neuropharmacology 33:387–391
Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (eds) (1993) Current protocols in molecular biology. Wiley, New York
Boess FG, Martin IL (1994) Molecular biology of 5-HT receptors. Neuropharmacology 33:275–317
Bradford M (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
Chanda PK, Minchin MCW, Davis AR, Greenberg L, Reilly Y, McGregor WH, Bhat R, Lubeck MD, Mizutani S, Hung PP (1993) Identification of residues important for ligand binding to the human 5-hydroxytryptamine1A serotonin receptor. Mol Pharmacol 43:516–520
Charest A, Wainer BH, Albert PR (1993) Cloning and differentiation-induced expression of a murine serotonin1A receptor in a septal cell line. J Neurosci 13:5164–5171
Erdmann J, Shimron-Abarbanell D, Cichon S, Albus M, Meier W, Lichtermann D, Minges J, Renner U, Franzek E, Ertl MA, Hebebrand J, Remschmidt H, Lehmkuhl G, Poustka F, Schmidt M, Fimmers R, Körner J, Rietschel M, Propping P, Nöthen MM (1994) Systematic screening for mutations in the promoter and the coding region of the 5-HT1A gene. Am J Med Genet 60:393–399
Fletcher A, Cliffe IA, Dourish CT (1993) Silent 5-HT1A receptor antagonists: utility as research tools and therapeutic agents. Trends Pharmacol Sci 14:441–448
Guan X-M, Peroutka SJ, Kobilka BK (1992) Identification of a single amino acid residue responsible for the binding of a class of β-adrenergic receptor antagonists to 5-hydroxytrypiamine1A receptors. Mol Pharmacol 41:695–698
Hoyer D, Pazos A, Probst A, Palacios JM (1986) Serotonin receptors in the human brain. I. Characterization and autoradiographic localization of 5-HT1A recognition sites. Apparent absence of 5-HT1B recognition sites. Brain Res 376:85–96
Hoyer D, Clarke DE, Fozard JR, Hartig PR, Martin GR, Mylecharane EJ, Saxena PR, Humphrey PPA (1994) VII. International Union of Pharmacology classification of receptors for 5-hydroxytryptamine (serotonin). Pharmacol Rev 46:111–229
Kobilka BK, Frielle T, Collins S, Yang-Feng T, Kobilka TS, Francke U, Lefkowitz RJ, Caron MG (1987) An intronless gene encoding a potential member of the family of receptors coupled to guanine nucleotide regulatory proteins. Nature 329:75–79
Metcalf MA, McGuffin RW, Hamblin MW (1992) Conversion of the human 5-HT1Dβ serotonin receptor to the rat 5-HT1B ligand-binding phenotype by THR355ASN site directed mutagenesis. Biochem Pharmacol 44:1917–1920
Oksenberg D, Masters SA, O'Dowd BF, Jin H, Havlik S, Peroutka SJ, Ashkenazi A (1992) A single amino acid difference confers major pharmacological variation between human and rodent 5-HT1B receptors. Nature 360:161–163
Parker EM, Grisel DA, Iben LG, Shapiro RA (1993) A single amino acid difference accounts for the pharmacological distinctions between the rat and human 5-hydroxytryptamine1B receptor. J Neurochem 60:380–383
Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467
Seyfried CA, Greiner HE, Haase AF (1989) Biochemical and functional studies on EMD 49980: a potent, selectively presynaptic D-2 dopamine agonist with actions on serotonin systems. Eur J Pharmacol 160:31–41
Taylor DP (1990) Serotonin agents in anxiety Ann NY Acad Sci 600:545–556
Van Tol HHM, Bunzow JR, Guan H-C, Sunahara RK, Seeman P, Niznik HB, Civelli O (1991) Cloning of the gene for a human dopamine D4 receptor with high affinity for the antipsychotic clozapine. Nature 350:610–614
Zifa E, Fillion G (1992) 5-Hydroxytryptamine receptors. Pharmacol Rev 44:401–458
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Brüss, M., Bühlen, M., Göthert, M. et al. Binding properties of the naturally occurring human 5-HT1A receptor variant with the Ile28Val substitution in the extracellular domain. Naunyn-Schmiedeberg's Arch Pharmacol 352, 455–458 (1995). https://doi.org/10.1007/BF00172786
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DOI: https://doi.org/10.1007/BF00172786