Behavioral deficits and dopaminergic degeneration following developmental iron and manganese exposures in NRXN/NLGN loss-of-function Caenorhabditis elegans

Neurexin ( NRXN ) and neuroligin ( NLGN ) genes encode synaptic adhesion proteins vital to synaptic structure and function and have been established as autism susceptibility genes. Increasing evidence suggests that developmental exposure to excess metals may interact with genetic susceptibility to influence neurodevelopmental outcomes. Iron (Fe) and manganese (Mn) are considered essential micronutrients necessary for brain development and brain physiological function; however, their dyshomeostasis due to excessive accumulation may disrupt normal neuronal homeostasis and promote neurotoxicity. Yet interactions of these metals with ASD-associated genetic variants remain unclear. Here, we investigated the effects of acute developmental exposure to Fe and Mn on dopamine (DA)-dependent behavioral phenotypes in nrx-1 and nlg-1 loss-of-function Caenorhabditis elegans . Age-synchronized larval stage-1 (L1) worms of the wild-type N2, nrx-1 , and nlg-1 deletion mutants were treated with either FeCl 2 or MnCl 2 for 1 h. We assessed worm survival, whole-body metal content, glutathione levels, and RONS. Further, we assessed DA-dependent behaviors and DAergic degeneration. Our results show variation in LD 50 , ranging from 1.28 mM to 2.7 mM for Fe and 7.5 mM to 12.29 mM for Mn, with the nrx-1 strain showing the lowest LD50 in both paradigms. There was a significant increase in bioaccumulation of Fe and Mn in all strains following exposure. The nrx-1 strain showed a dose-dependent increase in GSSG, while all strains showed an increase in RONS post-exposure to Fe. There was a reduction in the number of body bends off food in the N2 strain, while the nrx-1 and nlg-1 strains displayed an increased ratio of body bends, also for basal slowing response and ethanol avoidance response after exposure to Mn and Fe. Furthermore, fluorescence imaging showed structural alteration in DAergic neurons following both metal exposures. Our study reveals gene-environmental interactions between ASD-associated genes NRXN and NLGN and metal exposure during development, with potential impacts on normal behavior and on oxidative stress and dopaminergic homeostasis.