<p>Dental enamel formation depends on tightly regulated mineral deposition and organic matrix removal during the maturation stage. Fluoride (F) adversely affects the maturation stage of amelogenesis, causing hypomineralized enamel defects. However, hypomineralized enamel defects have increased worldwide in recent years, making it urgent to understand how co-exposure to other environmental contaminants affects fluorosis and maturation-stage amelogenesis at fluoride levels sufficient to induce defects during early-life exposure conditions in experimental models. This study aimed to determine whether lead (Pb) co-exposure amplifies fluoride-induced disturbances in the enamel maturation stage by mapping depth-dependent changes in mineral, organic, and water components across the enamel layer. Pregnant Wistar rats and their offspring were exposed to F (50 ppm), Pb (30 ppm), or both in drinking water from gestation to postnatal day 30. Mandibular third molars were evaluated using a modified Thylstrup–Fejerskov index and analyzed by quantitative microradiography and polarized light microscopy to obtain depth-resolved compositional data. Mean defect scores were 0.0 (Control), 2.6 (F), and 3.2 (Pb + F). Entire enamel thickness mineral volume decreased from 71.0% (Control) to 41.1% (F) and to 35.2% (Pb + F), accompanied by a marked increase in organic matrix volume (9.3% to &gt; 30%) (<i>p</i> &lt; 0.0001). Spatial analyses showed minimal disturbances and smaller effect sizes at the superficial region, with progressively larger effects toward the central and inner enamel regions. These findings demonstrate that Pb co-exposure exacerbates fluoride-induced enamel hypomineralization and disrupts the normal spatial pattern of enamel maturation, emphasizing the importance of combined environmental exposures for understanding mechanisms governing the development and maturation of mineralized tissues.</p>