Vol. 56, Issue 1, 54-67, July 1999

Humanization of Mouse 5-Hydroxytryptamine_1B Receptor Gene by Homologous Recombination: In Vitro and In Vivo Characterization

P. Bonaventure,^{1 2} L. Umans,¹ M. H. M. Bakker, P. Cras, X. Langlois, W. H. M. L. Luyten, A. A. H. P. Megens, L. Serneels, F. Van Leuven, and J. E. Leysen

Department of Biochemical Pharmacology, Janssen Research Foundation, Beerse, Belgium (P.B., M.H.M.B., X.L., J.E.L.); Research Institute Neurosciences, Vrije Universiteit Amsterdam, the Netherlands (P.B., J.E.L.); Experimental Genetics Group, Centrum voor Menselijke Erfelijkheid, Vlaams Interuniversitair Instituut voor Biotechnologie, Katholieke Universiteit Leuven, Campus Gasthuisberg, Leuven, Belgium (L.U., L.S., F. Van L.); Department of Pathology, University of Antwerp, Edegem, Belgium (P.C.); Department of Functional Genomics, Janssen Research Foundation, Beerse, Belgium (W.H.M.L.L.); and Department of General In Vivo Pharmacology, Janssen Research Foundation, Beerse, Belgium (A.A.H.P.M.)

We replaced the coding region of the murine 5-hydroxytryptamine (5-HT)_1B receptor by the human 5-HT_1B receptor using homologous recombination in embryonic stem cells and generated and characterized homozygous transgenic mice that express only the human (h) 5-HT_1B receptor. The distribution patterns of h5-HT_1B and murine (m) 5-HT_1B receptor mRNA and binding sites in brain sections of transgenic and wild-type mice were identical as measured by in situ hybridization histochemistry and radioligand receptor autoradiography. When measured in parallel under identical conditions, the h5-HT_1B receptor expressed in mouse brain had the same pharmacological characteristics as that in human brain. Stimulation by 5-HT_1B agonists of [³⁵S]guanosine-5'-O-(3-thio)triphosphate binding in brain sections demonstrated the functional coupling of the h5-HT_1B receptor to G proteins in mouse brain. In tissue slices from various brain regions, electrically stimulated [³H]5-HT release was not modified by 5-HT_1B agonists in tissue from either transgenic and wild-type mice; a 5-HT_1B antagonist enhanced electrically stimulated [³H]5-HT release in wild-type mouse brain, but was ineffective in the transgenics. The centrally active 5-HT_1A/5-HT_1B agonist RU24969 induced hypothermia but did not increase locomotor activity in the transgenic mice. The ineffectiveness of RU24969 in the transgenic mice could be due to the lower affinity of the compound for the h5-HT_1B receptor compared with the m5-HT_1B receptor. The present study demonstrates a complete replacement of the mouse receptor by its human receptor homolog and a functional coupling to G proteins. However, modulation of [³H]5-HT release could not be shown. Furthermore, behavioral effects were not clearly observed, which may be due to a lack of appropriate tools.