(174a) Electro-Plasmonic Neural Stimulation and Its Implication for Prosthetic Devices

Authors: Ratka Damnjanovic^a,c, Parveen Bazard^a,c, Robert D. Frisina^a,b,c and Venkat R. Bhethanabotla^a,c

^a Dept. of Chemical and Biomedical Eng., ^b Dept. of Communication Sciences and Disorders, ^c Global Center for Hearing and Speech Research

University of South Florida, Tampa, FL –33620

Introduction:

Sensorineural prosthetic/therapeutic devices like cochlear implants, cardiac pacemakers and sciatic nerve stimulators, used for neural stimulation, are unable to achieve sub-micron level spatial resolution due to the spread of electric currents to the surrounding tissue around the stimulation points. Recently, nanoparticles-based optical stimulations have attracted considerable attention. Our previous study demonstrated the use of visible light for stimulation of SH-SY5Y neuroblastoma cell lines utilizing gold nanoparticles coated microelectrodes (Bazard et al., Sci. Rep. – 2017). This study expands the technique further by stimulating primary neurons with a hybrid – optical & electrical stimuli and therefore presents a more robust & novel stimulation for neurons, which is tunable to achieve the desired cellular responses.

Methods:

Microelectrode Development:

Microelectrodes were prepared as reported in the previous article (Bazard et al., Sci. Rep. – 2017). Briefly, glass micropipette was silanized with 10 % solution of γ-aminopropyl triethoxy silane in ethanol. Then, the tip of the micropipette was coated with 20 nm-diameter colloidal gold nanoparticles (Au NPs).

In-vitro Stimulations:

5-7 weeks C57B1/6 mice were used for the study. Mouse was decapitated, and the trigeminal ganglia nerve was removed. The nerve tissue was dissociated enzymatically, with HBSS containing collagenase type I and dispase II, and finally neurons were cultured in L15 media containing 10 % FBS and left to incubate in an incubator maintained at 37 C, 5 % CO₂ over night after dissociation. All cells were used within 36 hours timeframe.

All the electrophysiology experiments were done using whole cell patch clamp methods. Microelectrode was placed next to a single patched neuron cell and a 532 nm green laser beam was focused onto the gold coated electrode tip using an optical fiber. Whole cell patch-clamp techniques were used in conjunction with an Axopatch 700B amplifier, Digidata 1440 interface, and pCLAMP-9 software (Axon Instruments, Union City, CA, USA). Extracellular solution was used to flood the cells in the petri dish. For the hybrid stimulation, electrical stimulus was added, in addition to the optical plasmonic stimulation. Neural responses were recorded using the patch electrode.

Results:

Electrical stimulation was used before and after the optical stimulation to verify that cells are healthy and can fire action potentials. Initially, experiments were done with pure optical stimulations (1-5 ms pulse, 100-120 mW power). Plasmonic stimulations were more detrimental to the cells as compared to pure electrical stimulation. The success rate of firing action potential was very low, less than 20 % cells fired the action potential. Also, pure optical stimulations led to membrane alteration transiently. To overcome these issues, we combined electrical and optical stimulations to add the advantages from both stimulation methods, plasmonic and electrical, while decreasing the disadvantages from both methods when used alone. The percentage reduction in input current with hybrid stimulation, as compared to the current required to fire action potential with pure electrical stimulation, was approximately 40-50%. Also, electrical action potentials were recorded after hybrid stimulation at a higher success rate as compared to after pure plasmonic stimulation. We were also able to achieve multiple consecutive action potentials firings when applying consecutive hybrid stimulation on the cells. Further optimization of the lead and lag time of the electrical vs. plasmonic stimulus was also studied. The most optimal output is achieved when electrical stimulus leads before optical by 0.4 to 1.4 ms and when optical stimulus leads electrical by 0.6ms or less. The best repeatability of the action potential firing was achieved when electrical leads optical by less than 1 ms.

Summary:

After optimizing in-vitro stimulation parameters, our long-term goal is to develop nanoparticle-light based cochlear implants for the hearing-impaired population. The reduction of current required to trigger action potentials, and the evidence that cells stay healthy after repeated exposure to hybrid stimulation are ground breaking results for a new generation of tunable cochlear implants that can offer more precise frequency modulation enabled by more selective and tunable activation of auditory neurons.

Reference:

Bazard et al., Nanoparticles based Plasmonic Transduction for Modulation of Electrically Excitable Cells, Scientific Reports, 7 (7803), 2017