A great deal is now known about the chemical nature of prions and the mechanism by which they propagate. In contrast, how abnormal forms of the prion protein (PrP) kill nerve cells is still a mystery. There is evidence that the neurotoxicity of prions lies in their ability to alter or subvert a normal, physiological function of PrPC, the cellular form of PrP, but the details of this process are obscure. Tg(?CR) mice, which express a mutant PrP deleted for residues 105-125, provide powerful insights into prion related pathogenic mechanisms. These animals spontaneously develop a severe neurodegenerative illness that is reversed in a dose-dependent fashion by co-expression of wild-type PrP. We have been interested in elucidating the cellular and molecular mechanisms underlying the powerful toxicity of PrP?CR. We have recently discovered that the ?CR deletion acts as a dominant, gain-of-function mutation that strongly activates an ion channel activity that is intrinsic to, or is indirectly induced by, PrP. Moreover, we have found that disease-associated point mutations in the central region of PrP have a similar effect, suggesting that some familial prion diseases are due to excitotoxic activation of ion channels. In this application, we propose to critically test an "ion channel hypothesis" of prion diseases. First, we will survey all known human mutations in the central region of PrP for their effect on ion channel activity in vitro. We will then undertake characterization of the biophysical properties of the channels induced by PrP molecules carrying point and deletion mutations in the central region. Finally, we will determine if excitotoxic activation of ionotropic glutamate receptors plays a role in the neuronal death and neuropathology induced by mutant PrP molecules and infectious PrPSc. The pathogenic mechanisms elucidated in this project are likely to have wide applicability, since abnormal activation of ion channels is a well established paradigm in a number of other neurodegenerative diseases and animal models. Moreover, identification of specific ion channel targets for PrP-mediated toxicity would represent the first step in development of an entirely new class of anti-prion drugs that inhibit cellular neurotoxic pathways, rather than PrPSc propagation. PUBLIC HEALTH RELEVANCE: Prion diseases are fatal neurodegenerative disorders of humans and animals that pose a grave threat to public health, and endanger the safety of the food, blood and organ supplies. This application will explore a novel mechanism of prion pathogenesis based on abnormalities in the ion channels that control electrical activity in the nervous system. This project represents the first step in development of an entirely new class of anti-prion drugs that inhibit cellular neurotoxic pathways, rather than prion propagation.