Effect of in vivo chronic exposure to clotrimazole on zebrafish testis function

Clotrimazole is an azole fungicide used as a humanpharmaceutical that is known to inhibit cytochrome P450(CYP) enzymatic activities, including several steroidogenicCYP. In a previous report, we showed that a 7-day exposureto clotrimazole induced the expression of genes related tosteroidogenesis in the testes as a compensatory response, in-volving the activation of the Fsh/Fshr pathway. In this context,the aim of the present study was to assess the effect of an invivo 21-day chronic exposure to clotrimazole (30–197μg/L)on zebrafish testis function, i.e., spermatogenesis and androgenrelease. The experimental design combined (1) gene transcriptlevels measurements along the brain–pituitary–gonad axis, (2)11-ketotestosterone (11-KT) quantification in the blood, and(3) histology of the testes, including morphometric analysis.The chronic exposure led to an induction of steroidogenesis-related genes andfshrin the testes as well asfshβin thepituitary. Moreover, increases of the gonadosomatic indexand of the volume proportion of interstitial Leydig cells wereobserved in clotrimazole-exposed fish. In accordance withthese histological observations, the circulating concentrationof 11-KT had increased. Morphometric analysis of the testesdid not show an effect of clotrimazole on meiotic(spermatocytes) or postmeiotic (spermatids and spermatozoa)stages, but we observed an increase in the number of type Aspermatogonia, in agreement with an increase in mRNA levelsofpiwil1, a specific molecular marker of type A spermatogo-nia. Our study demonstrated that clotrimazole is able to affecttesticular physiology and raised further concern about theimpact of clotrimazole on reproduction