Fig. 1

Analysis of the conditional knockout mice with specific deletion of either MOR or DOR in primary afferent Na_v1.8-expressing neurons. A The scheme shows the Oprm1^fl conditional allele or “floxed” allele (Oprm1^fl/fl) with exons 2 and 3 of the MOR gene flanked by two loxP sites and the excised allele (deletion of the exons 2 and 3) resulting from the expression of Cre recombinase under the control of the Na_v1.8 sodium channel (SCN10A) gene promoter. Arrows indicate primers used to detect gene excision (A and B) and floxed allele (C and D) by PCR. PCR experiments show exon 2–3 deletion in DRGs but not in brain of MOR cKO mice. The two bands in DRGs result from gene excision in primary afferent nociceptive neurons (Na_v1.8⁺ neurons) but not in other (Na_v1.8-negative) cells. mRNA expression levels of Na_v1.8 (SCN10A) (B), MOR (Oprm1) (C) and DOR (Oprd1) (D) were quantified by quantitative RT-PCR in DRGs from MOR cKO mice (black histogram) and their wild-type MOR flox littermates (white histogram). The absence of non-specific germline activation of the Cre recombinase was confirmed by quantifying MOR (Oprm1) mRNA level in brain samples (E). mRNA expression levels were normalized to wild-type MOR flox littermates (n = 5–6 mice/group). F The scheme shows the Oprd1^fl conditional allele or “floxed” allele (Oprd1^fl/fl) with exon 2 of the DOR gene flanked by two loxP sites and the excised allele (exon 2 deletion) resulting from the expression of Cre recombinase under the control of the Na_v1.8 (SCN10A) gene promoter. Arrows indicate primers (E and F) used to examine gene excision by PCR. PCR depicts exon 2 deletion in genomic DNA from DRGs but not brain of DOR cKO mice. The two bands in DRGs result from gene excision in primary afferent nociceptive neurons (Na_v1.8⁺ neurons) but not in other (Na_v1.8-negative) cells. mRNA expression levels of Na_v1.8 (SCN10A) (G), DOR (Oprd1) (H) and MOR (Oprm1) (I) were quantified by quantitative RT-PCR in DRGs from DOR cKO mice (black histogram) and their wild-type DOR flox littermates (white histogram). The absence of non-specific germline activation of the Cre recombinase was confirmed by quantifying DOR (Oprd1) mRNA level in brain samples (J). mRNA expression levels were normalized to wild-type DOR flox littermates (n = 5–6 mice/group). Data are expressed as mean ± SEM. Statistical analysis was performed using Mann–Whitney U test. *p < 0.05, **p < 0.01 compared to wild-type littermate mice