Kynurenine pathway mediated multi-generational neurotoxicity induced by fluoride exposure in Caenorhabditis elegans.

Introduction

Global exposure to high-fluoride groundwater endangers approximately 200 million people, leading to endemic fluorosis in regions like India, Iran, and parts of China [1], [2], [3]. Beyond the well-established hydrogeological route, atmospheric transport (e.g., via sand and dust storms in arid regions) constitutes a significant complementary exposure pathway. Wang et al. [4], These multi-route exposures—through drinking water, inhaled dust, and fluoride-rich brick-tea—create a complex and persistent environmental challenge [5], [6], [7].

Fluoride is recognized as a developmental neurotoxicant. Epidemiological studies consistently associate early-life exposure with cognitive deficits and reduced IQ in children, a concern exacerbated by fluoride’s ability to cross the placental and blood-brain barriers [8], [9], [10], [11], [12], [13], [14]. In the ELEMENT cohort in Mexico, a negative correlation was found between maternal urinary fluoride concentration and offspring’s cognitive scores during childhood [15]; the MIREC cohort in Canada further confirmed the association between prenatal fluoride exposure and reduced IQ scores in offspring, especially in boys, within the North American context [16]; while a meta-analysis of studies conducted in high fluoride areas in China suggested the risk of intellectual impairment under long-term multigenerational exposure [17]. The children in these studies typically experienced long-term exposure from birth, which to some extent reflects the multigenerational exposure background of their families and environments. However, it is impossible to distinguish whether this exposure is directly attributed to the children themselves or to the residual effects of exposure from their mothers/grandparents. Evidence from mammalian models further suggests a potential intergenerational dimension to its neurotoxicity, with prenatal or multi-generational exposure leading to impaired neurodevelopment and progressive cognitive decline [18], [19]. However, the molecular mechanisms underlying these effects, particularly their persistence across generations, remain poorly understood [20].

These observations raise a pivotal question: what are the inherited molecular footprints that mediate such multi-generational effects? The nematode Caenorhabditis elegans provides a powerful model to dissect multi-generational toxicity, given its established utility in documenting rapid, non-genetic inheritance of stress responses from diverse environmental insults [21], [22], [23], [24], [25], [26], [27]. Such inheritance often involves mechanisms like the transmission of mitochondrial stress [28]. Despite the significant morphological differences between nematodes and humans, many key cellular and molecular pathways are highly conserved across evolution, including stress response pathways (such as the insulin/IGF-1 signaling pathway and MAPK pathway); neuronal development and function (such as synaptic formation, neurotransmitter synthesis, and transport); and epigenetic regulatory mechanisms (such as histone modification, non-coding RNA, and DNA methylation). These are the core mediators through which transgenerational effects occur.

A plausible mechanistic entry point lies in fluoride-induced disruption of central metabolic pathways. Emerging evidence hints at a specific interaction between fluoride and tryptophan metabolism, particularly the kynurenine pathway (KP). Geeraerts et al. [29], [30], This is highly significant because the KP, a major route of TRP catabolism, produces several neuroactive metabolites. Notably, kynurenic acid (KYNA), an endogenous N-methyl-D-aspartate receptor (NMDAR) antagonist, has been identified in C. elegans as a critical contributor to age-related cognitive decline [31]. The broader neurotoxicity of KP metabolites, often driven by inflammatory activation leading to oxidative stress and mitochondrial dysfunction, is well-documented in neurodegenerative contexts [32], [33].

We therefore hypothesize that fluoride exposure triggers persistent dysregulation of the kynurenine pathway, and that this dysregulation is a key mechanistic event underlying its observed multi-generational neurotoxicity. The KP, initiated by IDO/TDO enzymes, branches into distinct neuroactive arms: one leading to the potentially neurotoxic 3-HK and QUIN via KMO, and another to the NMDAR-antagonist KYNA via KAT [34], [35](Fig. 1). The imbalance of these metabolites represents a compelling, yet unproven, mechanism for fluoride-induced cognitive impairment across generations.

This study is designed to systematically test this hypothesis using C. elegans. We aim to determine whether multi-generational neurobehavioral deficits induced by NaF are mediated by specific alterations in the kynurenine pathway, thereby linking a fundamental metabolic disturbance to the inheritance of neurotoxicity. Elucidating this mechanism in C. elegans contributes to our understanding of fluoride’s long-term hazards and may inform future studies on metabolic pathways in other organisms.