Acute exposure to bis(2-ethylhexyl)phthalate disrupts calcium homeostasis, energy metabolism and induces oxidative stress in the testis of Danio rerio

Bis(2-ethylhexyl)phthalate (BEHP) negatively affects testicular functions in different animal species, disturbing reproductive physiology and male fertility. The present study investigated the in vitro acute effect of BEHP on the mechanism of action of ionic calcium (Ca²⁺) homeostasis and energy metabolism. In addition, the effect of BEHP on oxidative stress was studied in vitro and in vivo in the testis of Danio rerio (D. rerio). Testes were treated in vitro for 30 min with 1 μM BEHP for ⁴⁵Ca²⁺ influx measurements. Testes were also incubated with 1 μM BEHP for 1 h (in vitro) or 12 h (in vivo) for the measurements of lactate content, ¹⁴C-deoxy-d-glucose uptake, lactate dehydrogenase (LDH) and gamma-glutamyl transpeptidase (GGT) activity, total reactive oxygen species (ROS) production and lipid peroxidation. In addition, the effect of BEHP (1 μM) on GGT, glutamic oxaloacetic transferase (GOT) and glutamic pyruvic transferase (GPT) activity in the liver was evaluated after in vivo treatment for 12 h. BEHP disturbs the Ca²⁺ balance in the testis when given acutely in vitro. BEHP stimulated Ca²⁺ influx occurs through L-type voltage-dependent Ca²⁺ channels (L-VDCC), transitory receptor potential vaniloid (TRPV1) channels, reverse-mode Na⁺/Ca²⁺ exchanger (NCX) activation and inhibition of sarco/endoplasmic reticulum Ca²⁺-ATPase (SERCA). BEHP affected energy metabolism in the testis by decreasing the lactate content and LDH activity. In vitro and in vivo acute effects of BEHP promoted oxidative stress by increasing ROS production, lipid peroxidation and GGT activity in the testis. Additionally, BEHP caused liver damage by increasing GPT activity.