Ultralarge Modulation of Fluorescence by Neuromodulators in Carbon Nanotubes Functionalized with Self-Assembled Oligonucleotide Rings

Abstract

Noncovalent 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, which results 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 relative change in fluorescence intensity (Δ F/ F₀) of up to 3500%, a signal appropriate for in vivo neuroimaging, and uncover the photophysical principles and intermolecular interactions that govern the molecular recognition and fluorescence modulation of this nanosensor synthesized from the spontaneous self-assembly of (GT)₆ ssDNA rings 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 set of parameters with which to tune nanosensor dynamic range, analyte selectivity 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 rings around (9,4) SWNTs, inducing periodic surface potentials that modulate exciton recombination lifetimes. Further evidence is presented to elucidate how dopamine analyte binding modulates SWNT fluorescence. We discuss the implications of our findings for SWNT-based molecular imaging applications.

Keywords: Single wall carbon nanotubes; molecular dynamics simulations; molecular sensing; neuromodulation.

Figure 1.

Nanosensor response and selectivity for neuromodulators dopamine and norepinephrine as a function of polymer length (a, b) Near-infrared fluorescence spectra of (GT)₆-SWNT and (GT)₁₅-SWNT suspensions before (red trace) and after (black trace) addition of 100 μM dopamine (DA). Mean traces and standard deviation bands from n=3 measurements are presented. (c, d) Neurotransmitter analyte library chemical structure and heat map of ΔF/F₀ screen against (GT)_N-SWNT library. Analyte abbreviations: Ach = acetylcholine, 5-HT = serotonin, DA = dopamine, Hist = histamine, GABA = γ-aminobutyric acid, Glu = glutamate, Gly = glycine, Asp = aspartate, NE = norepinephrine. Heat map ΔF/F₀ are computed for the peak intensity of the (9,4) SWNT chirality (~1127 nm center wavelength) from the convoluted spectra and all measurements were made at pH~7. (e) ΔF/F₀ of each sequence suspension, for each SWNT chirality: (8,3) dark blue, (6,5) blue, (7,5) cyan, (10,2) green, (9,4) and (7,6) yellow, (8,6) and (12,1) red, (10,3) and (10,5) maroon. Insert: Baseline fluorescence intensity of (GT)_N suspensions of the (9,4) chirality (red trace) and change in its fluorescence intensity after addition of 100 μM of dopamine (orange trace). (f) (GT)₆-SWNT nanosensor response curve for norepinephrine (red) and dopamine (black) computed for the (9,4) SWNT peak intensity. Error bars are standard deviation from n = 3 independent measurements. Experimental data (circles) were fit with Hill equation (solid line).

Figure 2.

Solvatochromic shifts reveal neuromodulator-specific molecular interactions with nanosensors dependent on ssDNA sequence and length (a) Middle row: sodium cholate (SC) binds to exposed SWNT surfaces and displaces bound (GT)_N polymers. Bottom row: Nanosensor incubation in dopamine (DA) or norepinephrine (NE) stabilizes ssDNA polymers on the SWNT surface, disallowing SC from accessing the SWNT surface. Top row: Incubation in p-tyramine (TY) does not stabilize surface adsorbed ssDNA against displacement by SC (b) 1 wt.% SC induces a solvatochromic shift in SWNT fluorescence. The shift for the (GT)₆-SWNT conjugate is presented here as an example. (c) Fluorescence peak shift corresponding to the (9,4) SWNT chirality (~1127 nm) upon exposure to 1 wt.% SC without (dash trace) and with (solid trace) pre-incubation in 10 μM DA. Error bars are standard deviation from n = 3 measurements. Negative peak shits correspond to blue shifting of the peak in the emission spectrum, as shown in (b). (d) Time-resolved fluorescence measurements of (GT)₆-SWNT incubated in 10 μM DA (red trace), 10 μM p-tyramine (TY) (blue trace), and incubated in neither (orange trace). Upon addition of 0.25 wt. % SC indicated by the black bar, solvatochromic peak shift in the dopamine incubated corona is eliminated. (e) SC induced solvatochromic peak shift in (GA)₆-SWNT incubated in 10 μM of dopamine suggests (GA)₆ exhibits short lived stability on SWNT following dopamine incubation.

Figure 3.

Computational modeling of ssDNA-SWNT nanosensor complexes. (a) Representative conformation of (GT)₁₅-SWNT. SWNT is depicted as a gray surface, (GT)₁₅ and its backbone are shown in licorice and black ribbon representations, and ssDNA atoms are shown in gray (C), red (O), blue (N), and orange (P). (b) Representative conformation of (GT)₆-SWNT, containing three (GT)₆ polymers. The color scheme is the same as in panel a. (c) Electrostatic potential energy profile at the SWNT surface in the (GT)₁₅-SWNT system as a function of SWNT axial length. The profile is averaged over 2 ns and over the radial SWNT dimension, and includes the effects of the complete SWNT environment present in MD simulations (ssDNA, water, and ions). (d) Electrostatic potential energy profile at the SWNT surface for the (GT)₆-SWNT system plotted as a function of SWNT axial length. (e) Free energy landscape of (GT)₆-SWNT at 300 K. The structures corresponding to two free energy minima are labeled by indices 1 and 2. (f) Net charges of molecular fragments in the (GT)₂-SWNT system, evaluated in quantum mechanical calculations. (g) Net charges of molecular fragments in the (GT)₂-SWNT system with an adsorbed dopamine molecule, evaluated in quantum mechanical calculations. The color scheme in panels f and g: black (DNA), silver (non-terminal SWNT atoms), blue surface (terminal -CH groups capping the SWNT), yellow (sodium ions), green, blue, red and white spheres (C, N, O and H atoms on dopamine). (h) Electron (red) and hole (blue) probability densities in a Kronig-Penney potential (Methods). Probability density values are labeled on the left axis, and the values associated with the potential energy well are labeled on the right axis.