Investigating Potential Ryanodine Receptor Agonism of Environmental Pollutants Having Two Non-Coplanar Aromatic Rings

Abstract

Ryanodine receptors (RyRs) are a class of intracellular calcium channels present in neurons, cardiac muscle, and skeletal muscle which are thought to cause the release of calcium from the endoplasmic or sarcoplasmic reticulum upon activation by any one of several possible endogenous compounds. There is evidence to suggest that RyRs play a very important role in mediating a number of neurodevelopmental processes in both the central nervous system and the peripheral nervous system, and defects in RyR function may play a role in the development of some cases of autism and Alzheimer’s disease. Two organic pollutants, polychlorinated biphenyls (PCBs) and triclosan (TCS) have been shown to have a significant impact on calcium regulation via RyR activation in laboratory tests, and another, bisphenol A (BPA), is suspected of operating in a similar manner. The results of these studies have led some researchers, notably Pessah, Cherednichenko, & Lein (2010), to speculate that these results may provide evidence of a link between the increasing use of neurotoxic substances in agriculture, manufacturing, and personal care products and the shocking increases in neurodevelopmental disorders over the last several decades. This experiment aims to expand on this research by examining the impact of BPA on RyR function, and to investigate whether exposure to multiple agonists has a compounding effect on intracellular calcium regulation in vitro. To achieve this, cells of the murine myoblast cell line C2C12 were grown, differentiated into RyR-expressing myotubes, and exposed to various stimuli. Intracellular calcium concentration was monitored using a fluorescent assay before, during, and after treatment with 1μM of the selected stimulus. The results of this analysis showed that compared to the blank control, fluorescence values increased significantly (p < .05) in cells exposed to the control agonist caffeine and in those cells exposed to TCS and caffeine in combination. These results indicate that BPA does not show significantly increased calcium-release inducing activity at low levels of exposure but may show RyR agonist activity in more concentrated amounts, and that exposure to TCS in addition to other RyR agonists in a short period of time may have a compounding effect on RyR-mediated calcium release.