Identification of the functional mechanisms of alcohol–Wnt signaling pathway interactions in taste sensory organ development: Insight from the mexican tetra (Astyanax mexicanus)

Abstract

Animal models provide critical insight into conserved mechanisms underlying development and disease. The Mexican tetra (Astyanax mexicanus), which exists as both surface and cave morphs, offers a powerful system to study craniofacial and sensory development. Canonical Wingless/Integrated beta-catenin (Wnt/β-catenin) signaling is an important cell signaling pathway in early embryonic development and its disruption is implicated in various birth defects. Prenatal alcohol exposure (PAE) has been consistently associated with characteristic birth defects, categorized as Fetal Alcohol Spectrum Disorders (FASD). Alcohol exposure during embryogenesis suppresses both canonical and non-canonical Wnt signaling, associated with craniofacial abnormalities, while folic acid has been shown to minimise the early developmental defects. This study examined how alcohol exposure affects taste bud formation and gustatory associated chemoattractive behaviour, and whether modulation of Wnt signaling or folic acid supplementation alters these outcomes in surface fish (SF) and cavefish (CF) morphs. Embryos were exposed to ethanol (EtOH), Lithium chloride (LiCl) (Wnt activator), Wnt-C59 (WC-59) (Wnt inhibitor), or folic acid 10 hours post-fertilization (hpf). Taste bud patterning and distribution was assessed across larval stages at 8, 15 and 30 days post-fertilization (dpf) using live whole mount Ponceau S stain. The chemosensory attractive behaviour was assessed using the various taste stimulants in DanioVision observation system. CF consistently exhibited increased taste bud size and broader spatial distribution compared to SF, with divergence becoming more pronounced over development. Regional analysis revealed iii sequential emergence of taste bud subtypes, with T1, T3, and T4 appearing earlier, while T2 emerged later and was observed exclusively in CF within the stages examined. These findings suggest that differences in developmental timing and spatial organization underlie morph-specific sensory architectures. Across both surface and cave morphs of Mexican tetra, chemical perturbation of canonical Wnt signaling resulted in only limited and inconsistent effects on gustatory morphology. Although minor stage-specific differences were detected, these changes were not maintained across developmental stages or anatomical regions. Behavioral responses were similarly resilient to treatment, as chemoattractive behavior showed no consistent alterations in time spent near feed, distance to feed, or locomotor activity. Together, these findings suggest that modulation of canonical Wnt signaling has a minimal impact on taste bud development and feeding-related behavior in either morph under the conditions tested. Instead, behavioral outcomes were strongly morph-dependent: SF exhibited more variable and exploratory movement patterns, whereas CF demonstrated more consistent feed engagement and elevated locomotor activity. Overall, intrinsic differences between morphs were the dominant factor shaping both gustatory morphology and chemoattractive behaviour, indicating that these traits are robust to short-term developmental perturbation. These results suggest that sensory system organization in Astyanax mexicanus is primarily governed by developmental programming and evolutionary adaptation rather than acute modulation of signaling pathways under the conditions tested.