Sokolowski, D. | Search bioRxiv

bioRxiv2025

Methylphenidate exposure alters brain gene expression and induces transgenerational DNA methylation changes in Poecilia reticulata guppies

Chronic exposure to stimulants is known to affect behavioral phenotypes and epigenetic profiles intergenerationally. We have previously shown that chronic methylphenidate hydrochloride (MPH) exposure in male guppies (Poecilia reticulata) induces persistent, paternally transmitted behavioral changes across multiple unexposed generations. Here, we investigated the underlying epigenetic signatures of this transgenerational behavioral inheritance. Building on our previous study, which used composite behavioral scores, we investigates the transgenerational inheritance patters of individual behaviors and found a robust, male-lineage-dependent increase in swimming across unexposed G2-G4 female offspring. Our molecular analysis showed that that chronic MPH exposure in G1 males alters brain gene expression, with 76 differentially expressed genes including genes with developmental and locomotory functions, and an over-representation of long non-coding RNAs. Furthermore, we found that unexposed G4 descendants from MPH-treated lineages exhibit widespread changes in brain DNA methylation, including genes related to developmental and swimming behavior. Interestingly, we found one lnRNA, LOC103476631, as both differentially expressed in G1 males and differentially methylated in G4 fish, providing a compelling candidate for ncRNA-directed DNA methylation as a mechanism for the observed transgenerational epigenetic inheritance of swimming behavior. Author SummaryDrugs used to treat attention-deficit/hyperactivity disorder, such as methylphenidate, are widely prescribed to children and adolescents during key stages of brain development. We previously showed that exposing male guppies to a low, chronic dose of methylphenidate changed anxiety-like behavior not only in the treated fish but also in several generations of their unexposed descendants. In this study, we asked how such long-lasting behavioral effects might be recorded in the brain. We first confirmed that a simple measure of swimming behavior remains altered in descendants of treated males across multiple generations. We then examined brains from the directly exposed fathers and from great-grand-offspring that were never exposed to the drug. In fathers, we found changes in the activity of genes, including many long non-coding RNAs, a class of genes that has been linked to transgenerational inheritance. In great-grand-offspring, we found significantly altered DNA methylation profiles, an epigenetic mark that has also been associated with transgenerational inheritance. Together, our findings suggest that developmental exposure to a commonly used stimulant can leave a stable molecular memory in the brain that persists across generations.

genetics↗

bioRxiv2022

Postnatal developmental trajectory of sex-biased gene expression in the mouse pituitary gland

The pituitary gland regulates essential physiological processes such as growth, pubertal onset, stress response, metabolism, reproduction, and lactation. While sex biases in these functions and hormone production have been described, the underlying identity, temporal deployment, and cell-type specificity of sex-biased pituitary gene regulatory networks are not fully understood. To capture sex differences in pituitary gene regulation dynamics during postnatal development, we performed 3 untranslated region sequencing and small RNA sequencing to ascertain gene and microRNA expression respectively across five postnatal ages (postnatal days 12, 22, 27, 32, 37) that span the pubertal transition in female and male C57BL/6J mouse pituitaries (n=5-6 biological replicates for each sex at each age). We observed over 900 instances of sex-biased gene expression and 17 sex-biased microRNAs, with the majority of sex differences occurring with puberty. Using miRNA-gene target interaction databases, we identified 18 sex-biased genes that were putative targets of 5 sex-biased microRNAs. In addition, by combining our bulk RNA-seq with publicly available male and female mouse pituitary single-nuclei RNA-seq data, we obtained evidence that cell-type proportion sex differences exist prior to puberty and persist post-puberty for three major hormone-producing cell types: somatotropes, lactotropes, and gonadotropes. Finally, we predicted sex-biased genes in these three pituitary cell types after accounting for cell-type proportion differences between sexes. Our study reveals the identity and postnatal developmental trajectory of sex-biased gene expression in the mouse pituitary. This work also highlights the importance of considering sex biases in cell-type composition when understanding sex differences in the processes regulated by the pituitary gland. HighlightsO_LIMale and female mouse pituitary gland gene and miRNA expression was profiled across five postnatal ages spanning pubertal development C_LIO_LIAbundant sex differences in pituitary gene expression exist prior to puberty and become more prominent upon puberty C_LIO_LICombining expression data from genes and miRNAs revealed 18 putative sex-biased gene targets of 5 sex-biased miRNAs C_LIO_LISex differences in the proportions of somatotropes, lactotropes, and gonadotropes are predicted to occur prior to puberty C_LI

genomics↗