Scott Nawy - US grants

In the retina, the sign-inverting synapse between photoreceptors and ON bipolar cells is the foundation for the ON pathway in vision. The synapse is sign-inverting because glutamate, released in darkness by rod and cone photoreceptors, hyperpolarizes the membrane of ON bipolar cells. Hyperpolarization arises when binding of glutamate to the metabotropic receptor mGluR6 on the dendrites of the ON bipolar cell activates a G protein (Go), which then shuts off a synaptic transduction current, most likely through a membrane-delimited pathway. The channel, whose molecular identity is currently unknown, allows a mixture of cations to flow through it; as a result of this ion selectivity, the ON bipolar cell to depolarizes when light shuts off transmitter release from photoreceptors and allows these channels to open. Recent evidence from ours and other labs demonstrates that Ca2+ strongly inhibits the transduction current, contributing to the conversion of sustained to transient light responses in the retina. The goal of this proposal is to functionally identify the transduction channel(s) in mouse ON bipolar cells, and to elucidate the mechanism of regulation by Ca2+. The identity of the channel is currently unknown, but our recent findings suggest that it is a member of the TRP channel family in particular, and the vanilloid subgroup of the TRP family, TRPV1 in particular. TRPV1 agonist such as capsaicin and anandamide appear to open the ON bipolar cell transduction channel as well. Furthermore, TRPV1 channels are known to be regulated by a variety of intracellular messengers such as PIP2, Ca2+, and cAMP, many of which are also known to modulate the transduction channel in ON bipolar cells. Aims 1 and 2 of this proposal will examine the possibility that one or more TRPV1 agonist may serve as the endogenous activator of the transduction channel in bipolar cells of the mouse retina, as well as the mechanisms by which synthesis of these endogenous compounds are regulated. Aim III will follow up on our recent studies on Ca2+ regulation of transduction channels. The goal of this aim is to determine the mechanism by which Ca2+ depresses the transduction current, and it will also address the possibility that transduction channels expressed in rod and cone bipolar cells are differentially regulated by Ca2+. Results from these studies will provide insight into the fundamental mechanisms that regulate sensitivity and dynamic range in the visual system.

DESCRIPTION (provided by applicant): The rapid cycling of AMPA receptors (AMPARs) into and out of the membrane maintains neurotransmission at a number of CNS synapses. The cycling of AMPARs in the hippocampus and cortex is dynamically regulated by changes in levels of basal synaptic transmission and has been proposed to play a role in certain forms of synaptic plasticity. It remains unclear, however, whether the cycling of AMPARs also occurs at synapses not believed to exhibit postsynaptic forms of activity dependent plasticity. Additionally the question remains as to whether there are differences in the regulation of receptor trafficking at synapses that are subject to very different patterns of synaptic activation. For example, while many CNS synapses function primarily through intermittent, activity driven neurotransmitter release, synapses in the retina are subject to tonic glutamate release and stimulus dependent cessation of synaptic transmission. We are investigating the trafficking of AMPARs in retinal neurons and its regulation by activity. Our preliminary data demonstrates that, GluR2-containing AMPARs, can be rapidly cycled at the extrasynaptic membrane in the retina. However, contrary to in hippocampal synapses, activity in the retina stabilizes AMPARs in a non-cycling mode. This reversible process is modulated by physiological light stimuli. Experiments in this proposal, will seek to test the hypothesis, that normal light/dark cycles drive changes in the cycling of GluR2-containihg AMPARs thereby impacting functional signaling in the retina. Experiments in Aim 1 will establish the physiological conditions that mediate changes in the cycling of AMPARs in the retina. In Aim 2 we will seek to determine the molecular mechanisms by which activity links to changes in the cycling of AMPARs. Finally, in Aim 3 we will characterize the physiological significance of altered AMPAR cycling on the function of synaptic transmission in the retina. Results from these experiments will greatly enhance our understanding of the function and regulation .of signaling in the retina potentially identifying the existence a previously unknown form activity dependent of retinal plasticity. This should provide valuable insight into possible therapeutic treatments relevant to diseases of retinal development and degeneration.