| Ryanodine receptors (RyRs) and Inositol-1,4,5-trisphosphate receptors (IP3Rs) are channels that regulate Ca2+ release from the ER/SR of most cells, a process essential for many cellular functions. This dissertation focuses on the following factors affecting intracellular Ca2+ homeostasis: (1) Naturally occurring Xestospongin C (XeC) and its hydroxylated derivatives, (2) Malignant hyperthermia (MH) point mutations (MHRyR1), and (3) The effects of synthetic polychlorinated biphenyl 95 (PCB 95) on MHRyR1.; Although XeC has been shown to inhibit IP3Rs, its structural effects on RyR1 and SERCA have not been thoroughly examined. For the first time, direct measurements on SERCA activity revealed that XeC and its hydroxylated derivatives do not affect SERCA pumps. XeC potently inhibits IP3R and also RyR1, but at a much higher concentration. Similarly, its hydroxylated derivatives inhibit IP3R but activate RyR1 at the same concentrations. Thus, these hydroxylated derivatives of XeC become important biochemical tools for studying the involvement of RyR1 and IP3R in mobilizing intracellular Ca2+.; Genetic point mutations on RyR1 have been known to cause ME, a potentially fatal pharmacogenetic disorder of skeletal muscle. To understand the functional effects of seven selected MH point mutations, channel modulation by endogenous Ca2+ and Mg2+ were measured, which revealed that point mutations elevated resting calcium concentrations and also caused RyR1 to become hypersensitive toward Ca2+ activation and hyposensitive toward Mg2+ inhibition.; Recent data on synthetic PCBs has shown a relationship to significant morbidity in exposed humans. To understand how PCB 95 exposure to subjects possessing NM mutations affects the structure and function of RyR1, in vitro assays were performed using porcine skeletal muscle containing mutation R615C (analogous to human mutation R614C). Pre-existing functional effects of R615C on channel regulation by Ca2+ and Mg 2+ contribute significantly to activation of PCB 95 on MHRyR1 compared to that of WtRyR1. Although MH is a musculoskeletal disorder, the observed effects of PCB 95 on MHRyR1 suggest that PCB 95 would have similar negative effects in other organs that express RyR1. |
