Assessment of the Effects of Endocrine Disrupting Compounds on the Development of Vertebrate Neural Network Function Using Multi-electrode Arrays.

Bis-phenols, such as bis-phenol A (BPA) and bis-phenol-S (BPS), are polymerizing agents widely used in the production of plastics and numerous everyday products. They are classified as endocrine disrupting compounds (EDC) with estradiol-like properties. Long-term exposure to EDCs, even at low doses, has been linked with various health defects including cancer, behavioral disorders, and infertility, with greater vulnerability during early developmental periods. To study the effects of BPA on the development of neuronal function, we used an in vitro neuronal network derived from the early chick embryonic brain as a model. We found that exposure to BPA affected the development of network activity, specifically spiking activity and synchronization. A change in network activity is the crucial link between the molecular target of a drug or compound and its effect on behavioral outcome. Multi-electrode arrays are increasingly becoming useful tools to study the effects of drugs on network activity in vitro. There are several systems available in the market and, although there are variations in the number of electrodes, the type and quality of the electrode array and the analysis software, the basic underlying principles, and the data obtained is the same across the different systems. Although currently limited to analysis of two-dimensional in vitro cultures, these MEA systems are being improved to enable in vivo network activity in brain slices. Here, we provide a detailed protocol for embryonic exposure and recording neuronal network activity and synchrony, along with representative results.

Long-term Behavioral and Reproductive Consequences of Embryonic Exposure to Low-dose Toxicants.

Bisphenols, such as bisphenol A (BPA) and bisphenol S (BPS) are polymerizing agents widely used in the production of plastics and numerous everyday-use products. Based on their chemical structure and estradiol-like biological properties, they have been classified as endocrine disrupting compounds (EDC). Long-term exposure to EDCs, even at low doses, has been linked to various health defects including cancer, behavioral disorders and infertility, with greater vulnerability indicated during early developmental periods. Cellular and molecular studies with the genetically tractable nematode model Caenorhabditis elegans have demonstrated that exposure to BPA causes apoptosis, embryonic lethality and disruption in the DNA repair mechanisms. We have previously reported that exposure of C. elegans embryos to low doses of different bisphenols decreases fecundity. In addition, we have shown that the effects of exposure during the very early stages of development persist into adulthood as assayed by quantifying habituation behavior, a form of non-associative learning. Here, we provide detailed protocols for embryonic exposure to low-dose EDCs as well as the associated fecundity and anterior touch habituation assays, along with representative results.

Effects of BPA and BPS exposure limited to early embryogenesis persist to impair non-associative learning in adults.

BACKGROUND: Bisphenol-A (BPA) is a polymerizing agent used in plastic bottles and several routinely used consumer items. It is classified among endocrine disrupting chemicals suspected to cause adverse health effects in mammals ranging from infertility and cancer to behavioral disorders. Work with the invertebrate lab model Caenorhabditis elegans has shown that BPA affects germ cells by disrupting double-stranded DNA break repair mechanisms. The current study utilizes this model organism to provide insight into low-dose and long-term behavioral effects of BPA and bisphenol-S (BPS), a supposed safer replacement for BPA. FINDINGS: Experiments presented in our report demonstrate that the effects of embryonic exposure to considerably low levels of BPA persist into adulthood, affecting neural functionality as assayed by measuring habituation to mechano-sensory stimuli in C. elegans. These results are noteworthy in that they are based on low-dose exposures, following the rationale that subtler effects that may not be morphologically apparent are likely to be discernible through behavioral changes. In addition, we report that embryonic exposure to BPS follows a pattern similar to BPA. CONCLUSIONS: Building upon previous observations using the C. elegans model, we have shown that exposure of embryos to BPA and BPS affects their behavior as adults. These long-term effects are in line with recommended alternate low-dose chemical safety testing approaches. Our observation that the effects of BPS are similar to BPA is not unexpected, considering their structural similarity. This, to our knowledge, is the first reported behavioral study on low-dose toxicity of any endocrine disrupting chemical in C. elegans.