Benzodiazepines alter nucleotide and nucleoside hydrolysis in zebrafish (Danio rerio) brain

Abstract

Anxiety is characterized by unpleasant bodily sensations, such as pounding heart and intense fear. The therapy involves the administration of benzodiazepine drugs. Purinergic signaling participates in the induction of several behavioral patterns and their actions are inactivated by ectonucleotidases and adenosine deaminase (ADA). Since there is evidence about the involvement of purinergic system in the actions mediated by benzodiazepines, we evaluated the effects in vitro and in vivo of administration of diazepam and midazolam on nucleoside triphosphate diphosphohydrolases, ecto-5′-nucleotidase, and ADA activities in zebrafish brain, followed by the analysis of gene expression pattern of these enzymes and adenosine receptors (A1, A2a1, A2a2, A2b). The in vitro studies demonstrated that diazepam decreased ATP (66 % for 500 µM) and ADP hydrolysis (40–54 % for 10–500 µM, respectively). Midazolam decreased ATP (16–71 % for 10–500 µM, respectively) and ADP (48–73.5 % for 250–500 µM, respectively) hydrolysis as well as the ecto-ADA activity (26–27.5 % for 10–500 µM, respectively). AMP hydrolysis was decreased in animals treated with of 0.5 and 1 mg/L midazolam (32 and 36 %, respectively). Diazepam and midazolam decreased the ecto-ADA activity at 1.25 mg/L and 1 mg/L (31 and 33 %, respectively), but only 0.1 mg/L midazolam induced an increase (40 %) in cytosolic ADA. The gene expression analysis demonstrated changes on ecto-5′-nucleotidase, A1, A2a1, A2a2, and A2b mRNA transcript levels after acute treatment with benzodiazepines. These findings demonstrated that benzodiazepine exposure induces a modulation of extracellular nucleotide and nucleoside metabolism, suggesting the purinergic signaling may be, at least in part, related to benzodiazepine effects.