Our results indicate that abcb is

Our results indicate that abcb5 is likely to be a XT in zebrafish ionocytes, although, like renal cells, the phenotypes observed likely arise from the action of multiple transporters. Zebrafish lack an abcb1 ortholog, and abcb4 and abcb5 have been identified as the P-glycoproteins in zebrafish (Fischer et al., 2013). Efflux of RhB has been shown to be mediated by abcb4 which is ubiquitously expressed throughout the embryo (Fischer et al., 2013). Our results and those of the previous study suggest that the accumulation of ABC transporter substrates in ionocytes is not mediated by the same mechanism that handles RhB. Indeed, neither inhibitors nor gene knockdown shifted the accumulation of RhB from the yolk sac to the ionocytes (Fischer et al., 2013), indicating that the efflux mechanism for CAM, DiOC6, and BCECF-AM is distinct from that of RhB. abcb5 is a plausible transporter responsible for the efflux of ionocyteABC transporter substrates. In addition to its expression in the epidermal superficial stratum (Thisse and Thisse, 2004), our analysis of a recent single-cell RNA sequencing dataset shows that abcb5 is enriched in ionocytes of the 24 hpf embryo (Wagner et al., 2018; Fig. 7). Further studies will be needed tovalidate the functional role for abcb5 in this process, as well as other P-gp and MRP proteins present in ionocytes. Additionally, a recent study has demonstrated that zebrafish have OATs that can take up fluorescent small molecules (Dragojević et al., 2018), but the specific location and function of these transporters in the embryo remains to be determined. It is conceivable that one or more of these transporters, along with abcb5, is responsible for the uptake and efflux pattern observed in HR cells. Ionocytes have been previously compared to mammalian renal np e with regard to their role in ion homeostasis. The uptake and efflux activities we have observed in ionocytes suggest that ionocytes share additional functions and similarities with mammalian renal cells. Specifically, these additional functions could relate to detoxification, as mammalian renal cells are enriched with XTs, including P-gp and MRPs, as well as uptake transporters such as OATs (Giacomini et al., 2010). The differences among ionocyte subtypes are also reminiscent of the mammalian kidney, which exhibits XT, and ion transport, expression differences between renal cell types (Peng et al., 1999; Kojima et al., 2002; Huls et al., 2008). The overarching implication of this study is that zebrafish ionocytes may play an important role in toxicant defense for the zebrafish embryo. Though the data presented in this study suggest a plausible role for ionocytes in toxicant defense, the exact physiological implication of xenobiotic transporter function in zebrafish ionocytes remains unclear. A role for ionocytes in protection from xenobiotics aligns well with the conclusion of previous studies that have also suggested a role for these cells in embryo protection. For instance, when exposed to silver nanoparticles, enhanced gstp expression was observed to colocalize with ionocytes (Osborne et al., 2016), and glutathione conjugates are major substrates for ABCC transporters (Deeley et al., 2006; Slot et al., 2011; Luckenbach et al., 2014). Being one of the few cell types directly exposed to the external environment during embryonic development, ionocytes could represent a direct pathway for small, hydrophobic molecules to be removed from the embryo and effluxed into the environment. When internal barrier tissues are not yet fully functional, ionocytes may behave like renal cells to protect zebrafish embryonic development. However, ionocyte XTs could also be used to efflux an endogenous metabolite(s), protect ionocytes themselves, or act to reduce overall xenobiotic concentrations. Further studies will be needed to characterize the “transportome” of the ionocytes and distinguish among their functional roles.