Fortunately, we were able to administer these non-absorbed antibiotics via the drinking water in a manner that allowed the animals to maintain normal fluid intake and body weight, but significantly depleted the intestinal microbiota (Fig. 1). Importantly, these effects are independent of potential effects of antibiotic treatment on cocaine metabolism. Given that cocaine is primarily metabolized by the liver, it was important for our purposes to know that the alterations of gut microbiota did not affect the metabolism of cocaine – which could have affected the effective dose of cocaine each animal received. When we assayed for serum benzoylecygonine, the primary cocaine metabolite that is present in the blood, in a time course after high or low dose cocaine doses we saw no differences in control and antibiotic-treated animals (Fig. 2a,b) – suggesting that our strategy to reduce gut microbiota did not affect cocaine metabolism through either direct or indirect mechanisms. Similarly, the dramatic reduction in gut bacteria could have potentially resulted in alterations in the HPA axis which could have had confounding effects on our behavioral models. However, when
