Biology subjects

Beyene, A. G.

Publications and source records attributed to Beyene, A. G..

3 recordsLinked to original sources

bioRxiv2018

Imaging Striatal Dopamine Release Using a Non-Genetically Encoded Near-Infrared Fluorescent Catecholamine Nanosensor

Neuromodulation plays a critical role in brain function in both health and disease. New optical tools, and their validation in biological tissues, are needed that can image neuromodulation with high spatial and temporal resolution, which will add an important new dimension of information to neuroscience research. Here, we demonstrate the use of a catecholamine nanosensor with fluorescent emission in the 1000-1300 nm near-infrared window to measure dopamine transmission in ex vivo brain slices. These near-infrared catecholamine nanosensors (nIRCats) represent a broader class of nanosensors that can be synthesized from non-covalent conjugation of single wall carbon nanotubes (SWNT) with single strand oligonucleotides. We show that nIRCats can be used to detect catecholamine efflux in brain tissue driven by both electrical stimulation or optogenetic stimulation. Spatial analysis of electrically-evoked signals revealed dynamic regions of interest approximately 2 microns in size in which transients scaled with simulation intensity. Optogenetic stimulation of dopaminergic terminals produced similar transients, whereas optogenetic stimulation of glutamatergic terminals showed no effect on nIRCat signal. Bath application of nomifensine prolonged nIRCat fluorescence signal, consistent with reuptake blockade of dopamine. We further show that the chemically synthetic molecular recognition elements of nIRCats permit measurement of dopamine dynamics in the presence of dopamine receptor agonists and antagonists. These nIRCat nanosensors may be advantageous for future use because i) they do not require virus delivery, gene delivery, or protein expression, ii) their near-infrared fluorescence facilitates imaging in optically scattering brain tissue and is compatible for use in conjunction with other optical neuroscience tool sets, iii) the broad availability of unique near-infrared colors have the potential for simultaneous detection of multiple neurochemical signals, and iv) they are compatible with pharmacology. Together, these data suggest nIRCats and other nanosensors of this class can serve as versatile new optical tools to report dynamics of extracellular neuromodulation in the brain.

neuroscience↗

bioRxiv2018

Ultralarge Modulation of Single Wall Carbon Nanotube Fluorescence Mediated by Neuromodulators Adsorbed on Arrays of Oligonucleotide Rings

Non-covalent interactions between single-stranded DNA (ssDNA) oligonucleotides and single wall carbon nanotubes (SWNTs) have provided a unique class of tunable chemistries for a variety of applications. However, mechanistic insight into both the photophysical and intermolecular phenomena underlying their utility is lacking, resulting in obligate heuristic approaches for producing ssDNA-SWNT based technologies. In this work, we present an ultrasensitive \"turn-on\" nanosensor for neuromodulators dopamine and norepinephrine with strong {Delta}F/F0 of up to 3500%, a signal appropriate for in vivo imaging, and uncover the photophysical principles and intermolecular interactions that govern the molecular recognition and fluorescence modulation of this nanosensor synthesized from the non-covalent conjugation of (GT)6 ssDNA strands on SWNTs. The fluorescence modulation of the ssDNA-SWNT conjugate is shown to exhibit remarkable sensitivity to the ssDNA sequence chemistry, length, and surface density, providing a wealth of parameters with which to tune nanosensor dynamic range and strength of fluorescence turn-on. We employ classical and quantum mechanical molecular dynamics simulations to rationalize our experimental findings. Calculations show that (GT)6 ssDNA form ordered loops around SWNT, inducing periodic surface potentials that modulate exciton recombination lifetimes. Further evidence is presented to elucidate how analyte binding modulates SWNT fluorescence. We discuss the implications of our findings for SWNT-based molecular sensing applications.

neuroscience↗

bioRxiv2017

Molecular Recognition of Dopamine with Dual Near Infrared Excitation-Emission Two-Photon Microscopy

A key limitation for achieving deep imaging in biological structures lies in photon absorption and scattering leading to attenuation of fluorescence. In particular, neurotransmitter imaging is challenging in the biologically-relevant context of the intact brain, for which photons must traverse the cranium, skin and bone. Thus, fluorescence imaging is limited to the surface cortical layers of the brain, only achievable with craniotomy. Herein, we describe optimal excitation and emission wavelengths for through-cranium imaging, and demonstrate that near-infrared emissive nanosensors can be photoexcited using a two-photon 1560 nm excitation source. Dopamine-sensitive nanosensors can undergo two-photon excitation, and provide chirality-dependent responses selective for dopamine with fluorescent turn-on responses varying between 20% and 350%. We further calculate the two-photon absorption cross-section and quantum yield of dopamine nanosensors, and confirm a two-photon power law relationship for the nanosensor excitation process. Finally, we show improved image quality of the nanosensors embedded 2 mm deep into a brain-mimetic tissue phantom, whereby one-photon excitation yields 42% scattering, in contrast to 4% scattering when the same object is imaged under two-photon excitation. Our approach overcomes traditional limitations in deep-tissue fluorescence microscopy, and can enable neurotransmitter imaging in the biologically-relevant milieu of the intact and living brain.

neuroscience↗