Toward Plasmonic Neural Probes: SERS Detection of Neurotransmitters through Gold-Nanoislands-Decorated Tapered Optical Fibers with Sub-10 nm Gaps.

Integration of plasmonic nanostructures with fiber-optics-based neural probes enables label-free detection of molecular fingerprints via surface-enhanced Raman spectroscopy (SERS), and it represents a fascinating technological horizon to investigate brain function. However, developing neuroplasmonic probes that can interface with deep brain regions with minimal invasiveness while providing the sensitivity to detect biomolecular signatures in a physiological environment is challenging, in particular because the same waveguide must be employed for both delivering excitation light and collecting the resulting scattered photons. Here, a SERS-active neural probe based on a tapered optical fiber (TF) decorated with gold nanoislands (NIs) that can detect neurotransmitters down to the micromolar range is presented. To do this, a novel, nonplanar repeated dewetting technique to fabricate gold NIs with sub-10 nm gaps, uniformly distributed on the wide (square millimeter scale in surface area), highly curved surface of TF is developed. It is experimentally and numerically shown that the amplified broadband near-field enhancement of the high-density NIs layer allows for achieving a limit of detection in aqueous solution of 10-7  m for rhodamine 6G and 10-5  m for serotonin and dopamine through SERS at near-infrared wavelengths. The NIs-TF technology is envisioned as a first step toward the unexplored frontier of in vivo label-free plasmonic neural interfaces.

Tapered fibertrodes for optoelectrical neural interfacing in small brain volumes with reduced artefacts.

Deciphering the neural patterns underlying brain functions is essential to understanding how neurons are organized into networks. This deciphering has been greatly facilitated by optogenetics and its combination with optoelectronic devices to control neural activity with millisecond temporal resolution and cell type specificity. However, targeting small brain volumes causes photoelectric artefacts, in particular when light emission and recording sites are close to each other. We take advantage of the photonic properties of tapered fibres to develop integrated 'fibertrodes' able to optically activate small brain volumes with abated photoelectric noise. Electrodes are positioned very close to light emitting points by non-planar microfabrication, with angled light emission allowing the simultaneous optogenetic manipulation and electrical read-out of one to three neurons, with no photoelectric artefacts, in vivo. The unconventional implementation of two-photon polymerization on the curved taper edge enables the fabrication of recoding sites all around the implant, making fibertrodes a promising complement to planar microimplants.

Holographic Manipulation of Nanostructured Fiber Optics Enables Spatially-Resolved, Reconfigurable Optical Control of Plasmonic Local Field Enhancement and SERS.

Integration of plasmonic structures on step-index optical fibers is attracting interest for both applications and fundamental studies. However, the possibility to dynamically control the coupling between the guided light fields and the plasmonic resonances is hindered by the turbidity of light propagation in multimode fibers (MMFs). This pivotal point strongly limits the range of studies that can benefit from nanostructured fiber optics. Fortunately, harnessing the interaction between plasmonic modes on the fiber tip and the full set of guided modes can bring this technology to a next generation progress. Here, the intrinsic wealth of information of guided modes is exploited to spatiotemporally control the plasmonic resonances of the coupled system. This concept is shown by employing dynamic phase modulation to structure both the response of plasmonic MMFs on the plasmonic facet and their response in the corresponding Fourier plane, achieving spatial selective field enhancement and direct control of the probe's work point in the dispersion diagram. Such a conceptual leap would transform the biomedical applications of holographic endoscopic imaging by integrating new sensing and manipulation capabilities.

An open source three-mirror laser scanning holographic two-photon lithography system.

Two-photon polymerization is a widely adopted technique for direct fabrication of 3D and 2D structures with sub-diffraction-limit features. Here we present an open-hardware, open-software custom design for a holographic multibeam two-photon polymerization system based on a phase-only spatial light modulator and a three-mirror scanhead. The use of three reflective surfaces, two of which scanning the phase-modulated image along the same axis, allows to overcome the loss of virtual conjugation within the large galvanometric mirrors pair needed to accommodate the holographic projection. This extends the writing field of view among which the hologram can be employed for multi-beam two-photon polymerization by a factor of ~2 on one axis (i.e. from ~200µm to ~400µm), with a voxel size of ~250nm × ~1050nm (lateral × axial size), and writing speed of three simultaneous beams of 2000 voxels/s, making our system a powerful and reliable tool for advanced micro and nano-fabrications on large area.

Orthogonalization of far-field detection in tapered optical fibers for depth-selective fiber photometry in brain tissue.

The field of implantable optical neural interfaces has recently enabled the interrogation of neural circuitry with both cell-type specificity and spatial resolution in sub-cortical structures of the mouse brain. This generated the need to integrate multiple optical channels within the same implantable device, motivating the requirement of multiplexing and demultiplexing techniques. In this article, we present an orthogonalization method of the far-field space to introduce mode-division demultiplexing for collecting fluorescence from the implantable tapered optical fibers. This is achieved by exploiting the correlation between the transversal wavevector k t of the guided light and the position of the fluorescent sources along the implant, an intrinsic property of the taper waveguide. On these bases, we define a basis of orthogonal vectors in the Fourier space, each of which is associated with a depth along the taper, to simultaneously detect and demultiplex the collected signal when the probe is implanted in fixed mouse brain tissue. Our approach complements the existing multiplexing techniques used in silicon-based photonics probes with the advantage of a significant simplification of the probe itself.

Influence of the anatomical features of different brain regions on the spatial localization of fiber photometry signals.

Fiber photometry is widely used in neuroscience labs for in vivo detection of functional fluorescence from optical indicators of neuronal activity with a simple optical fiber. The fiber is commonly placed next to the region of interest to both excite and collect the fluorescence signal. However, the path of both excitation and fluorescence photons is altered by the uneven optical properties of the brain, due to local variation of the refractive index, different cellular types, densities and shapes. Nonetheless, the effect of the local anatomy on the actual shape and extent of the volume of tissue that interfaces with the fiber has received little attention so far. To fill this gap, we measured the size and shape of fiber photometry efficiency field in the primary motor and somatosensory cortex, in the hippocampus and in the striatum of the mouse brain, highlighting how their substructures determine the detected signal and the depth at which photons can be mined. Importantly, we show that the information on the spatial expression of the fluorescent probes alone is not sufficient to account for the contribution of local subregions to the overall collected signal, and it must be combined with the optical properties of the tissue adjacent to the fiber tip.

Flexible SAW Microfluidic Devices as Wearable pH Sensors Based on ZnO Nanoparticles.

In this work, a new flexible and biocompatible microfluidic pH sensor based on surface acoustic waves (SAWs) is presented. The device consists of polyethylene naphthalate (PEN) as a flexible substrate on which aluminum nitride (AlN) has been deposited as a piezoelectric material. The fabrication of suitable interdigitated transducers (IDTs) generates Lamb waves (L-SAW) with a center frequency ≈500 MHz traveling in the active region. A SU-8 microfluidics employing ZnO nanoparticles (NPs) functionalization as a pH-sensitive layer is fabricated between the IDTs, causing a shift in the L-SAW resonance frequency as a function of the change in pH values. The obtained sensitivity of ≈30 kHz/pH from pH 7 to pH 2 demonstrates the high potential of flexible SAW devices to be used in the measurement of pH in fluids and biosensing.

Comparative study of autofluorescence in flat and tapered optical fibers towards application in depth-resolved fluorescence lifetime photometry in brain tissue.

As the scientific community seeks efficient optical neural interfaces with sub-cortical structures of the mouse brain, a wide set of technologies and methods is being developed to monitor cellular events through fluorescence signals generated by genetically encoded molecules. Among these technologies, tapered optical fibers (TFs) take advantage of the modal properties of narrowing waveguides to enable both depth-resolved and wide-volume light collection from scattering tissue, with minimized invasiveness with respect to standard flat fiber stubs (FFs). However, light guided in patch cords as well as in FFs and TFs can result in autofluorescence (AF) signal, which can act as a source of time-variable noise and limit their application to probe fluorescence lifetime in vivo. In this work, we compare the AF signal of FFs and TFs, highlighting the influence of the cladding composition on AF generation. We show that the autofluorescence signal generated in TFs has a peculiar coupling pattern with guided modes, and that far-field detection can be exploited to separate functional fluorescence from AF. On these bases, we provide evidence that TFs can be employed to implement depth-resolved fluorescence lifetime photometry, potentially enabling the extraction of a new set of information from deep brain regions, as time-correlating single photon counting starts to be applied in freely-moving animals to monitor the intracellular biochemical state of neurons.

Investigation of the exciton relaxation processes in poly(9,9-dioctylfluorene-co-benzothiadiazole):CsPbI1.5Br1.5 nanocrystal hybrid polymer-perovskite nanocrystal blend.

The combination of lead halide perovskite nanocrystals and conjugated polymer in a blend film opens the way to the realization of hybrid active layers with widely tunable optical and electrical properties. However, the interaction between the polymeric and the perovskite component of the blends is mainly unexplored to date. In this work we perform temperature-dependent photoluminescence and time resolved photoluminescence measurements in order to deeply investigate the photophysics of a poly(9,9-dioctylfluorene-co-benzothiadiazole) (F8BT):CsPbI1.5Br1.5 nanocrystal hybrid film. Our results suggest that the primary interaction channel is charge transfer, both from F8BT to the NCs and from the NCs to F8BT, while Förster resonant energy transfer has no visible effects. Moreover, we show that the charge transfer is assisted by energy migration within the F8BT excited state distribution and that it is dependent on the local micromorphology of the film. Our work improves the current understanding of the polymer:perovskite NC interactions in hybrid films, and it is expected to be relevant for the development of hybrid organic-perovskite optoelectronic devices.

Two-photon fluorescence-assisted laser ablation of non-planar metal surfaces: fabrication of optical apertures on tapered fibers for optical neural interfaces.

We propose a feedback-assisted direct laser writing method to perform laser ablation of fiber optic devices in which their light-collection signal is used to optimize their properties. A femtosecond-pulsed laser beam is used to ablate a metal coating deposited around a tapered optical fiber, employed to show the suitability of the approach to pattern devices with a small radius of curvature. During processing, the same pulses generate two-photon fluorescence in the surrounding environment and the signal is monitored to identify different patterning regimes over time through spectral analysis. The employed fs beam mostly interacts with the metal coating, leaving almost intact the underlying silica and enabling fluorescence to couple with a specific subset of guided modes, as verified by far-field analysis. Although the method is described here for tapered optical fibers used to obtain efficient light collection in the field of optical neural interfaces, it can be easily extended to other waveguide-based devices and represents a general approach to support the implementation of a closed-loop laser ablation system of fiber optics.

High transmission from 2D periodic plasmonic finite arrays with sub-20 nm gaps realized with Ga focused ion beam milling.

Fabricating plasmonic nanostructures with good optical performances often requires lengthy and challenging patterning processes that can hardly be transferred to unconventional substrates, such as optical fiber tips or curved surfaces. Here we investigate the use of a single Ga focused ion beam process to fabricate 2D arrays of gold nanoplatelets for nanophotonic applications. While observing that focused ion beam milling of crossing tapered grooves inherently produces gaps below 20 nm, we provide experimental and theoretical evidence for the spectral features of grooves terminating with a sharp air gap. We show that transmission near 10% can be obtained via two-dimensional nano-focusing in a finite subset of 2D arrays of gold nanoplatelets. This enables the application of our nanostructure to detect variations in the refractive index of thin films using either reflected or transmitted light when a small number of elements are engaged.

Ray tracing models for estimating light collection properties of microstructured tapered optical fibers for optical neural interfaces.

Tapered optical fibers (TFs) were recently employed for depth-resolved monitoring of functional fluorescence in subcortical brain structures, enabling light collection from groups of a few cells through small optical windows located on the taper edge [Pisano et al., Nat. Methods16, 1185 (2019)1548-709110.1038/s41592-019-0581-x]. Here we present a numerical model to estimate light collection properties of microstructured TFs implanted in scattering brain tissue. Ray tracing coupled with the Henyey-Greenstein scattering model enables the estimation of both light collection and fluorescence excitation fields in three dimensions, whose combination is employed to retrieve the volume of tissue probed by the device.

Depth-resolved fiber photometry with a single tapered optical fiber implant.

Fiber photometry is increasingly utilized to monitor fluorescent sensors of neural activity in the brain. However, most implementations are based on flat-cleaved optical fibers that can only interface with shallow tissue volumes adjacent to the fiber. We exploit modal properties of tapered optical fibers (TFs) to enable light collection over an extent of up to 2 mm of tissue and multisite photometry along the taper. Using a single TF, we simultaneously observed distinct dopamine transients in dorsal and ventral striatum in freely moving mice performing a simple, operant conditioning task. Collection volumes from TFs can also be engineered in both shape and size by microstructuring the nonplanar surface of the taper, to optically target multiple sites not only in the deep brain but, in general, in any biological system or organ in which light collection is beneficial but challenging because of light scattering and absorption.

The Three-Dimensional Signal Collection Field for Fiber Photometry in Brain Tissue.

Fiber photometry is used to monitor signals from fluorescent indicators in genetically-defined neural populations in behaving animals. Recently, fiber photometry has rapidly expanded and it now provides researchers with increasingly powerful means to record neural dynamics and neuromodulatory action. However, it is not clear how to select the optimal fiber optic given the constraints and goals of a particular experiment. Here, using combined confocal/2-photon microscope, we quantitatively characterize the fluorescence collection properties of various optical fibers in brain tissue. We show that the fiber size plays a major role in defining the volume of the optically sampled brain region, whereas numerical aperture impacts the total amount of collected signal and, marginally, the shape and size of the collection volume. We show that ~80% of the effective signal arises from 105 to 106 µm3 volume extending ~200 µm from the fiber facet for 200 µm core optical fibers. Together with analytical and ray tracing collection maps, our results reveal the light collection properties of different optical fibers in brain tissue, allowing for an accurate selection of the fibers for photometry and helping for a more precise interpretation of measurements in terms of sampled volume.