Alleviation of specific responses in the combined exposure of freshwater mussel Unio tumidus to psychoactive substances and microplastics.

The environmentally relevant aquatic pollution is associated with the mixtures of xenobiotics, each in the low, picomolar to micromolar concentrations. Among these substances, the combinations of pharmaceuticals and microplastics (MP) have become an increasingly serious threat. The objective of this study was to track the specific and multi-stress responses of swollen river mussels (Unio tumidus) to the psychoactive substances caffeine (Caff) and chlorpromazine (Cpz) under combined exposure with MP. The MP (1 mg·L, size 35-50 μm), Caff (20 µg·L), Cpz (12 ng·L) or their mixture (Mix) were administered to mussels for 14 days. The redox state, enzymes of biotransformation and apoptosis were analysed in the digestive gland. All exposures except Mix caused oxidative injury to lipids and proteins, accompanied by increased GSH and metallothionein levels, suppressed NAD and activation of GST (except Mix), and GTPase. MP had the lower particular impact. Specific responses to Caff were activation of Cyp450 (EROD) and cathepsin D, decreased GSH/GSSG ratio and prominent demetallation of metallothionein. The Cpz caused an increase in NADH/NAD ratio and caspase-3 inhibition. In the combined exposure, the specific responses to single xenobiotics were alleviated which was confirmed by discriminant analysis. The Mix-group was distinguished by the highest NADH/NAD and GSH/GSSG ratios, markedly increased caspase-3 activity accompanied by the decrease of protein carbonyl level and the highest IBR index, attesting to the negative cumulative effect of multi-stress exposure. The vulnerability of mussels to pM concentration of neuroleptic Cpz needs particular attention.