The aim of the present study was to investigate the mechanisms underlying the desensitization of 5-HT_1Areceptors in the dorsal raphe and hypothalamus of serotonin (5-HT) transporter knock-out mice (5-HTT −/−). The density of 5-HT_1Areceptors in the dorsal raphe was reduced in both male and female 5-HTT −/− mice. This reduction was more extensive in female than in male 5-HTT −/− mice. 8-OH-DPAT-induced hypothermia was absent in female 5-HTT −/− and markedly attenuated in 5-HTT +/− mice. The density of 5-HT_1Areceptors also was decreased significantly in several nuclei of the hypothalamus, amygdala, and septum of female 5-HTT −/− mice. 5-HT_1Areceptor mRNA was reduced significantly in the dorsal raphe region, but not in the hypothalamus or hippocampus, of female 5-HTT +/− and 5-HTT −/− mice. G-protein coupling to 5-HT_1Areceptors and G-protein levels in most brain regions were not reduced significantly, except that G_oand G_i1proteins were reduced modestly in the midbrain of 5-HTT −/− mice. These data suggest that the desensitization of 5-HT_1Areceptors in 5-HTT −/− mice may be attributable to a reduction in the density of 5-HT_1Areceptors. This reduction is brain region-specific and more extensive in the female mice. The reduction in the density of 5-HT_1Areceptors may be mediated partly by reduction in the gene expression of 5-HT_1Areceptors in the dorsal raphe, but also by other mechanisms in the hypothalamus of 5-HTT −/− female mice. Finally, alterations in G-protein coupling to 5-HT_1Areceptors are unlikely to be involved in the desensitization of 5-HT_1Areceptors in 5-HTT −/− mice.