The overall goal of this research is to understand the relationship between ion channel modulation and neuronal function. Sodium channels are responsible for the initial depolarization of the action potential in most neurons. Because of this central role in neuronal activity, modulation of sodium channels can have powerful effects. Sodium channels are regulated by cAMP-dependent protein kinase and by protein kinase C. They are also modulated by direct activation of G proteins. Our preliminary data demonstrate that directly activating G proteins with guanine nucleotides or by expression of recombinant G protein subunits results in strong sodium channel modulation. Modulation of sodium channels by neurotransmitters acting on G-protein-coupled receptors linked to these pathways has been little studied. The experiments are designed to identify the G-protein coupled receptors that activate this modulation under physiological conditions and to characterize the biochemical pathways by which this G-protein-activation-dependent sodium channel modulation occurs. To identify neurotransmitters that activate these G-protein-dependent pathways under physiological conditions, a series of physiologically relevant neuromodulators will be tested in acutely isolated or primary cultured hippocampal neurons for effects that overlap those caused by direct activation of G proteins. Since sodium channel modulation by G protein activation occurs in the absence of exogenously-added ATP, the investigator will test the hypothesis that it is a membrane-delimited phenomenon that does not require protein phosphorylation. He will also test the hypothesis that it occurs by direct interaction of G protein subunits with the sodium channel as has been proposed for G protein modulation of muscarinic potassium channels and voltage-dependent calcium channels. Sites on the sodium channel required for G protein regulation will be identified and the possibility of direct G protein binding to the sodium channel will be addressed. These studies will be carried out using patch clamp recording from hippocampal neurons in combination with GTP analogs, toxins, antisense constructs and antibodies. Modulation will be reconstituted in mammalian tissue culture cells heterologously expressing, receptor, G protein subunits and rat brain sodium channels. Direct interaction between sodium channels and G protein subunits will be assessed biochemically. These studies will provide a careful assessment of the strength and potency of sodium channel modulation in brain cells. In addition, they will provide a detailed examination of the pathway by which sodium channel modulation does and can occur in this model brain system. Sodium channels are key players in all aspects of neuronal activity. An understanding of how they are modulated physiologically will provide the basis for developing pharmacologic means of controlling them to achieve specific therapeutic goals.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
First Independent Research Support & Transition (FIRST) Awards (R29)
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Physiology Study Section (PHY)
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Stewart, Randall
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University of Washington
Schools of Medicine
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Mantegazza, Massimo; Yu, Frank H; Powell, Andrew J et al. (2005) Molecular determinants for modulation of persistent sodium current by G-protein betagamma subunits. J Neurosci 25:3341-9
Mantegazza, M; Yu, F H; Catterall, W A et al. (2001) Role of the C-terminal domain in inactivation of brain and cardiac sodium channels. Proc Natl Acad Sci U S A 98:15348-53