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Single-Molecule Patch-Clamp FRET Imaging Microscopy in Living Cells

Award Number: R01GM098089
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 09/01/2012
PERIOD OF PERFORMANCE END DATE: 08/31/2018

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Single-Molecule Patch-Clamp FRET Imaging Microscopy in Living Cells - DESCRIPTION (provided by applicant): We propose to use single-molecule spectroscopy imaging simultaneously with single-channel current recording to interrogate the dynamics in the conformational motions of a single ion-channel, to better understand the mechanism of channel function at the cell membranes. To meet a major methodology and technical need and challenge in the medical life sciences, the primary goal of our proposal is to solve a critical problem that holds understanding ion channel protein biological function and dynamics over the last four decades ever since the patch-clamp electrophysiological technique demonstrated. Specifically, we propose to conduct a systematic technical development and demonstration. Our project consists of three primary aims: (1) Achieve an experimental understanding of the sub-unit conformational changes that control the open-close activity of the NMDA receptor; (2) Resolve the conformational changes of the NMDA receptor in desensitization and inactivation; and (3) Demonstrate high time resolution for single-molecule ion channel dynamics studies. Our proposed technical approach, patch-clamp confocal single-molecule fluorescence imaging microscopy, will be applicable for a broad range of ion channel proteins and receptors in various cells, including all of the cellular systems that have been traditionally studied by conventional patch-clamp electric current measurements and proteins that can be probed by fluorescence. We will use the N-methyl-D-aspartate (NMDA) receptor in living cells as a model system to demonstrate our innovative physical technique and its unprecedented applications in life sciences and medical researchers. The dynamics of NMDA receptor mediated calcium currents is crucial to the normal function of the brain. The temporal behavior of NMDA receptor activity is regulated by the dynamic conformational changes of the protein in response to agonist binding and dissociation. Identifying the conformational motions of the receptor is critical for understanding the mechanisms of receptor function. Inhomogeneous conformational motions of the receptor control the activation, inactivation, desensitization and deactivation of the channel that in turn shape the calcium transients.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2019 ( Subtotal = -$2,099 )
2019	2017	BOWLING GREEN STATE UNIVERSITY	319 ADMINISTRATION BLDG	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	001	4	3/4/2019	SUPPLEMENT FOR EXPANSION	-$2,099
2019	2015	BOWLING GREEN STATE UNIVERSITY	319 ADMINISTRATION BLDG	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	4	3/4/2019	NON-COMPETING CONTINUATION	$0
														Subtotal = -$2,099

Issue Date FY: 2017 ( Subtotal = $96,787 )
2017	2017	BOWLING GREEN STATE UNIVERSITY	319 ADMINISTRATION BLDG	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	001	4	9/20/2017	SUPPLEMENT FOR EXPANSION	$96,787
2017	2015	BOWLING GREEN STATE UNIVERSITY	319 ADMINISTRATION BLDG	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	4	8/17/2017	NON-COMPETING CONTINUATION	$0
														Subtotal = $96,787

Issue Date FY: 2015 ( Subtotal = $411,300 )
2015	2015	BOWLING GREEN STATE UNIVERSITY	***	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	3	6/26/2015	SUPPLEMENT FOR EXPANSION	$175,000
2015	2015	BOWLING GREEN STATE UNIVERSITY	***	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	001	4	8/20/2015	NON-COMPETING CONTINUATION	$236,300
														Subtotal = $411,300

Issue Date FY: 2014 ( Subtotal = $236,300 )
2014	2014	BOWLING GREEN STATE UNIVERSITY	***	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	3	8/26/2014	NON-COMPETING CONTINUATION	$236,300
														Subtotal = $236,300

Issue Date FY: 2013 ( Subtotal = $228,030 )
2013	2013	BOWLING GREEN STATE UNIVERSITY	106 University Hall	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	2	8/20/2013	NON-COMPETING CONTINUATION	$228,030
														Subtotal = $228,030

Issue Date FY: 2012 ( Subtotal = $236,300 )
2012	2012	BOWLING GREEN STATE UNIVERSITY	106 University Hall	BOWLING GREEN	OH	43403	WOOD	USA	Biomedical Research and Research Training	000	1	8/30/2012	NEW	$236,300
														Subtotal = $236,300

Grand Total All Awards = $1,206,618

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Single-Molecule Patch-Clamp FRET Imaging Microscopy in Living Cells

Award Number: R01GM098089

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 09/01/2012

PERIOD OF PERFORMANCE END DATE: 08/31/2018

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