C. elegans has been frequently used for studies on the mechanism of action, molecular targets, and behavioral and metabolic effects of fluoxetine, but data on the toxicokinetics of fluoxetine and its major metabolite norfluoxetine in C. elegans are lacking. In this study, we evaluated the toxicokinetics and relative potency of norfluoxetine and fluoxetine in chemotaxis and activity tests. Toxicokinetics experiments were conducted with varying times, concentrations of fluoxetine, and in the absence or presence of E. coli.
Methods
Effects of fluoxetine and norfluoxetine (racemic mixture) on C. elegans activity were tested. Worms were exposed in S medium with E. coli at an OD600 of 0.4 to nine concentrations of fluoxetine and norfluoxetine, ranging from 1 ng/L to 100 mg/L. The general activity was measured with a WMicrotracker from Phylumtech. The total number of interruptions over a 30 min period was calculated with the Wmicrotracker software. As C. elegans develop, their activity increases. To adjust for variations in the signal caused by the number of worms in each well, a baseline measurement of activity was taken after 50 h (the point at which a stable activity was reached). After this, worms were exposed and exposure-related changes in activity were quantified continuously for 24 h for each well specifically.
Results
Effects of fluoxetine and norfluoxetine on C. elegans activity were observed (Figure 2). The two compounds induced similar dose–response patterns, but a two-way ANOVA indicated significant effects of both concentration (p < 0.001) and compound (p = 0.0018). Only at 10 mg/L, norfluoxetine exposure resulted in a significantly lower activity index compared to fluoxetine (p < 0.01 with Bonferroni post hoc analysis). The respective EC50s (+95% confidence intervals) for the effect on activity were 40.0 (11.4–68.5) mg/L for fluoxetine and 10.1 (3.18–17.0) mg/L for norfluoxetine. So, there is an indication that norfluoxetine is slightly more potent.
Figure 2. Effect of fluoxetine and norfluoxetine on C. elegans (a) activity. Mean ± SEM is represented, and brackets with * indicate p < 0.05 between fluoxetine and norfluoxetine at that concentration level.
Environ. Sci. Technol. 2024. https://pubs.acs.org/doi/10.1021/acs.est.3c07744
Merel A. van der Most, Wouter Bakker, Sebastiaan Wesseling, and Nico W. van den Brink.