IADR Abstract Archives
Botulinum Toxin Inhibits SERCA1a, 2a and 2b in Temporalis Muscle.
Objectives: To test the effect of botulinum toxin type A on sarco-endoplasmic reticulum Calcium-ATPase (SERCA) isoforms 1a, 2a and 2b in temporalis muscle.
Methods: experimental research on SERCA isolated from rabbit temporalis muscles (n=12). Isoforms were isolated by differential ultracentrifugation and separated by high affinity chromatography. The enzymatic activity for each of the isoforms was assayed in vitro in standardized media containing 0.1 mg/mL SERCA, 0.1 mmol/L Ca-EGTA, 100 mMol/L KCL, 3 mmol/L MgCl2, 3 mmol/L ATP, 10 μmol/L calcimycin, and variable units (U) of botulinum toxin type A, at 37oC and pH 7.2. Pi production was determined by Fiske’s colorimetric method and taken as an index of the enzyme's activity. The inhibitory concentration (IC50) was determined (mean, SEM) and compared by ANOVA and SNK tests (p<0.05).
Results: botulinum toxin type A selectively inhibited the enzymatic activity of temporalis SERCA isoforms in a concentration-dependent manner. The inhibitory effect showed an inverse sigmoidal profile for the three of the isoforms. The IC50 was 11.0±2.1 U for SERCA2b, 18.1±1.2 for 2a and 32.1±3.0 for 1a. The most sensitive isoform to botulinum toxin was 2b (F=24.5, p<0.0001).
Conclusions: Since botulinum toxin inhibits SERCA isoforms in temporalis muscle at concentrations lower than currently recommended clinical doses (40 U), the inhibition of SERCA should also be considered as responsible for temporalis rigidity resulting from the therapeutic use of the drug.
Methods: experimental research on SERCA isolated from rabbit temporalis muscles (n=12). Isoforms were isolated by differential ultracentrifugation and separated by high affinity chromatography. The enzymatic activity for each of the isoforms was assayed in vitro in standardized media containing 0.1 mg/mL SERCA, 0.1 mmol/L Ca-EGTA, 100 mMol/L KCL, 3 mmol/L MgCl2, 3 mmol/L ATP, 10 μmol/L calcimycin, and variable units (U) of botulinum toxin type A, at 37oC and pH 7.2. Pi production was determined by Fiske’s colorimetric method and taken as an index of the enzyme's activity. The inhibitory concentration (IC50) was determined (mean, SEM) and compared by ANOVA and SNK tests (p<0.05).
Results: botulinum toxin type A selectively inhibited the enzymatic activity of temporalis SERCA isoforms in a concentration-dependent manner. The inhibitory effect showed an inverse sigmoidal profile for the three of the isoforms. The IC50 was 11.0±2.1 U for SERCA2b, 18.1±1.2 for 2a and 32.1±3.0 for 1a. The most sensitive isoform to botulinum toxin was 2b (F=24.5, p<0.0001).
Conclusions: Since botulinum toxin inhibits SERCA isoforms in temporalis muscle at concentrations lower than currently recommended clinical doses (40 U), the inhibition of SERCA should also be considered as responsible for temporalis rigidity resulting from the therapeutic use of the drug.
