Fig. 2. Sensitivity to halothane follows dRyr copy number. (A ) Reduced dRyr copy number is associated with resistance to halothane. Heterozygous dRyr deletion mutant dRyr δ25 /+ (medium gray ), carrying one copy of dRyr , is resistant to halothane compared with the control, Canton-S , which carries two copies. dRyr δ25 /dRyr GS21220 transheterozygotes (light gray ) are resistant compared with the wild-type and dRyr GS21220 /+ (dark gray ). (B ) The dRyr duplication strain dRyr 24D03 /+ (black ), carrying three copies of dRyr , is more sensitive to halothane than its control (VK33 without insertion, carrying two copies of dRyr ). Introducing dRyr 24D03 into insertion mutant dRyr GS21220 (dRyr GS21220 /+ ; dRyr 24D03 /+ , dark gray ) rescues the resistant phenotype of dRyr GS21220 /+ (gray ) to normal levels. Values are mean ± SEM. (C ) In the absence of anesthetic dRyr δ25 /+ (gray ), dRyr GS21220 /+ (dark gray ), and dRyr 24D03 (black ) display normal locomotor activity, whereas Ryr δ25 /dRyr GS21220 (light gray ) is significantly less active than control flies. Values are mean ± SEM (n = 3–5), and the asterisk denotes statistical significance (P = 0.0006, one-way ANOVA and Bonferroni–Dunn post hoc tests). (D ) dRyr copy number affects dRyr protein expression. (D1 ) Quantification of dRyr protein levels. Asterisks denote statistically significant difference from the wild-type (P < 0.0001 and P < 0.0001, one-way ANOVA and Bonferroni–Dunn post hoc tests). Values are mean ± SEM (n = 8). (D2 ) A representative Western blot, showing trend of increasing dRyr expression with the number of dRyr copies. dRyr = Drosophila Ryr .