Neurons form complex networks and communicate through synaptic connections. The molecular dynamics of cell surface molecules at synaptic terminals are essential for elucidating synaptic transmission and plasticity in biological neural networks. To achieve artificial control of synaptic transmission in neural networks at the single-synapse level, we propose and demonstrate the application of optical trapping for laser-induced perturbation to cellular molecules on neurons. In this study, we investigated the effects of optical forces on the dynamics of cell molecules in an optical trap on neurons. The diffusion properties of the cell surface molecules under optical trapping were evaluated using fluorescence analysis with single-particle tracking and fluorescence correlation spectroscopy. Molecular diffusion at the cell surface of neurons was compared to that of lipid molecules in artificial bilayers. Moreover, the molecular dynamics in an optical trap without fluorescent labeling under live cell conditions was evaluated using Raman spectroscopy.
Neuronal stimulation is essential to understand information processing in brain systems. Spatiotemporal patterns of neuronal activity can be modified by external stimuli. Recent studies have shown that neurons can be stimulated by short-pulse laser processing of the cell membrane. An optical vortex with a helical wavefront possesses an orbital angular momentum (OAM) enables the inward twisting of ablated materials, thereby processing further precisely cells beyond a conventional Gaussian beam. We herein study the mechanisms of neuronal stimulation with a focused nanosecond optical vortex. The focused nanosecond optical vortex on the cell membrane of rat hippocampal neurons induces extracellular Ca2+ influx and neuronal activity elicitation. Morphological changes of the neuronal cell membrane due to nanosecond optical vortex irradiation is also evaluated with fluorescence recovery after photobleaching. After the deposition of a single pulse of nanosecond optical vortex on the cell membrane of neurons, the fluorescence intensity of membrane probe at the focal region significantly decreases, however, it recovers within 5 seconds. Such dynamics suggests that the transient disruption occurs at the cell membrane based on laser ablation and recovers due to lateral diffusion of membrane molecules. The diffusion coefficients of membrane molecules after optical vortex irradiation are larger than those of Gaussian beam irradiation, and the disrupted membrane areas are smaller than the expected ones as the optical vortex focal region. These differences are attributed to the fact that the disruption of cell membrane owing to laser ablation and subsequent membrane diffusion are assisted by OAM transfer effects.
Neurons in the brain communicate by releasing and receiving neurotransmitters at synapse. Synaptic vesicles (SVs) that encapsulate neurotransmitters play an important role for neuronal communication. We demonstrate that optical trapping of synaptic vesicles in cultured rat hippocampal neurons regulates the neuronal network activity. The neuronal electrical activity was evaluated by extracellular potential measurement using microelectrodes arrays (MEAs). When a near-infrared trapping laser was focused on synaptic vesicles labeled with FM1-43 dye, fluorescence caused by two-photon absorption was observed at the focal spot. The fluorescence intensity gradually increased during the laser irradiation time at the laser power of 500 mW, indicating that optical trapping forces cause the assembly of SVs at the focal spot. In the extracellular potential measurement of neuronal electrical activity, spike number of spontaneous neuronal activity increased under optical trapping of SVs. The synchronicity of neuronal network activity by cross-correlation analysis increased after the laser irradiation under higher laser power conditions. These results suggest that neuronal electrical activity can be manipulated by optical trapping of synaptic vesicles.
For the purpose of precise manipulation of single nanoparticles by optical trapping, we demonstrated optical trapping of nanoparticles enhanced depending on the wavelength of excitation laser. The optical trapping dynamics of quantum dot (QD) nanoparticles at the focal spot was evaluated by fluorescence correlation spectroscopy (FCS). The simultaneous irradiation with excitation and near-infrared lasers increased the average transit time of QDs at the focal spot, which depended on the laser power and the wavelength of the excitation laser. This suggests that the particle motion of QD nanoparticles is constrained at the laser focus due to enhancement of optical trapping based on the resonant optical response.
We demonstrate surface plasmon resonance (SPR) based optical trapping of quantum-dot (QD) nanoparticles suspended
in water with a bull’s eye-type plasmonic chip. The particle dynamics of QD suspensions at the laser focus was evaluated
by fluorescence correlation spectroscopy. The average transit time of QD suspensions on the plasmonic chip increased than
that on the coverslip, suggesting that single QD was more constrained at the focal spot due to optical trapping enhanced
with SPR.
We numerically investigate the convection of surrounding fluid in optical trapping of micro- and nanoparticles. The
effects of the laser irradiation on the fluid simulation are twofold. First, we take into account the temperature increase of
the fluid due the photothermal effect of the solvent, that is, the fluid flow is described by the Navier-Stokes equations
under the Boussinesq approximation. Second, we assume that the suspended particles drag the fluid when they are
transported by the optical force. This dragging effect is considered in the fluid simulation by adding to the Navier-Stokes
equation an external forcing term, which is modelled by considering the counterbalance between the optical scattering
force and the Stokes drag. It is shown that the latter effect is dominant under the usual experimental setup in optical
trapping of particles with the diameter larger than 0.5 μm. Furthermore, the particle size dependence on the convective
flow speed is investigated. The numerical results are supported by optical trapping experiment qualitatively.
AMPA-type glutamate receptor (AMPAR) is one of neurotransmitter receptors at excitatory synapses in neuronal cell. For realizing the artificial control of synaptic transmission, we have applied optical trapping of quantum-dot (QD) conjugated AMPARs on neuronal cells. Here, we demonstrate simultaneous measurement combined with optical trapping and patch-clamp recordings to evaluate the neuronal electrical activity. The relationship between optical trapping dynamics of QD-AMPARs located on neuronal cells and the neuronal electrical activity was discussed.
AMPA-type glutamate receptor (AMPAR) is one of the major neurotransmitter receptors at excitatory synapses. The initial assembling states of AMPARs at cell surface are essential for synaptic transmission, which is related with learning and memory in living neural systems. To realize artificial control of synaptic transmission, we demonstrate to modulate the initial assembling states of quantum-dot conjugated AMPARs (QD-AMPARs) with optical trapping. The optical trapping dynamics of QD-AMPARs on living neurons was evaluated with fluorescence imaging and fluorescence correlation spectroscopy (FCS). The transit time at laser focus of QD-AMPARs on neurons estimated from FCS analysis increased with the culturing days and addition of neurotransmitter, which suggests that QD-AMPARs are confined at the focal spot due to optical trapping.
Molecular dynamics of glutamate receptor, which is major neurotransmitter receptor at excitatory synapse located on neuron, is essential for synaptic plasticity in the complex neuronal networks. Here we studied molecular dynamics in an optical trap of AMPA-type glutamate receptor (AMPAR) labeled with quantum-dot (QD) on living neuronal cells with fluorescence imaging and fluorescence correlation spectroscopy (FCS). When a 1064-nm laser beam for optical trapping was focused on QD-AMPARs located on neuronal cells, the fluorescence intensity of QD-AMPARs gradually increased at the focal spot. Using single-particle tracking of QD-AMPARs on neurons, the average diffusion coefficient decreased in an optical trap. Moreover, the decay time obtained from FCS analysis increased with the laser power and the initial assembling state of AMPARs depended on culturing day, suggesting that the motion of QD-AMPAR was constrained in an optical trap.
Molecular dynamics at synaptic terminals in neuronal cells is essential for synaptic plasticity and subsequent modulation
of cellular functions in a neuronal network. For realizing artificial control of living neuronal network, we demonstrate
laser-induced perturbation into molecular dynamics in the neuronal cells. The optical trapping of cellular molecules such
as synaptic vesicles or neural cell adhesion molecules labeled with quantum dots was evaluated by fluorescence imaging
and fluorescence correlation spectroscopy. The trapping and assembling dynamics was revealed that the molecular
motion was constrained at the focal spot of a focused laser beam due to optical trapping force. Our method has a
potential to manipulate synaptic transmission at single synapse level.
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