Voltage sensitive dyes (VSD) are used to record and image spatial and temporal patterns of electrical activity in cells and tissues. This laboratory ha been engaged in the design, synthesis and application of VSDs. The dyes that have emerged from this effort are widely used to optically record electrical signals where electrode-based measurements would be unsuitable. In this proposed continuation of our research program, we plan to develop better dyes and use them to elucidate a fundamental issue in synaptic signaling. We also will engage in a variety of collaborative projects that will span scales from the subcellular to the clinic in both neuronal and cardiac systems.
The first Aim proposes to build on the success of the new fluorinated VSDs with their improved photostability and sensitivity. We will modify these chromophores to adapt them to experiments that require different membrane binding, solubility and spectral properties to serve both our own experiments and those of our collaborators. Example collaborations include: marrying optical recording with optogenetic stimulation in hippocampal slices from transgenic mice;use of optical parametric oscillators for 2-photon excitation of long wavelength VSDs in brain slices and in vivo;merging optical recording with STED microscopy to image voltage in spines at nanometer resolution;imaging action potential propagation within single cardiomyocytes via rapid laser positioning;in vivo imaging of electrical activity in the heart;characterization of action potential propagation aroun lesions in human hearts.
The second Aim proposes to develop a completely novel mechanism for voltage sensing - "dipper dyes". As opposed to existing mechanisms which may produce at most a 60%/100mV change in fluorescence, the dipper dyes can in principle produce a several fold response to an action potential. In the third Aim, we will apply VSDs in combination with non-linear optical microscopy to investigate the voltage change at individual dendritic spines in response to neurotransmitter. These experiments, to be performed in mouse brain slices using 2-photon microscopy, will directly address the question of whether the spine can serve as an electrical compartment. We will also explore how electrical signals from pairs of neighboring spines can be integrated in the dendrite.
This project will develop new fluorescent voltage sensors that will permit the imaging of electrical activity in excitable tissue with sub-cellular resolution. Ths technology will be applied to the study of normal and diseased heart. It will also be used to understand information processing in the brain from the level of a synapse to an entire neural circuit.
|Acker, Corey D; Hoyos, Erika; Loew, Leslie M (2016) EPSPs Measured in Proximal Dendritic Spines of Cortical Pyramidal Neurons. eNeuro 3:|
|Crocini, C; Ferrantini, C; Scardigli, M et al. (2016) Novel insights on the relationship between T-tubular defects and contractile dysfunction in a mouse model of hypertrophic cardiomyopathy. J Mol Cell Cardiol 91:42-51|
|Crocini, Claudia; Coppini, Raffaele; Ferrantini, Cecilia et al. (2016) T-Tubular Electrical Defects Contribute to Blunted Î²-Adrenergic Response in Heart Failure. Int J Mol Sci 17:|
|Crocini, Claudia; Ferrantini, Cecilia; Coppini, Raffaele et al. (2016) Optogenetics design of mechanistically-based stimulation patterns for cardiac defibrillation. Sci Rep 6:35628|
|Frank, Pinar; Siebenhofer, Bernhard; Hanzer, Theresa et al. (2015) Proteo-lipobeads for the oriented encapsulation of membrane proteins. Soft Matter 11:2906-8|
|Loew, Leslie M (2015) Design and Use of Organic Voltage Sensitive Dyes. Adv Exp Med Biol 859:27-53|
|Brown, Sherry-Ann; McCullough, Louise D; Loew, Leslie M (2015) Computational neurobiology is a useful tool in translational neurology: the example of ataxia. Front Neurosci 9:1|
|Loew, Leslie M; Lewis, Aaron (2015) Second Harmonic Imaging of Membrane Potential. Adv Exp Med Biol 859:473-92|
|Wilson, Stacy A; Millard, Andrew; Lewis, Aaron et al. (2014) Monitoring membrane potential with second-harmonic generation. Cold Spring Harb Protoc 2014:643-54|
|Brown, Sherry-Ann; Loew, Leslie M (2014) Integration of modeling with experimental and clinical findings synthesizes and refines the central role of inositol 1,4,5-trisphosphate receptor 1 in spinocerebellar ataxia. Front Neurosci 8:453|
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