Conopeptide-Functionalized Nanoparticles Selectively Antagonize Extrasynaptic N-Methyl-d-aspartate Receptors and Protect Hippocampal Neurons from Excitotoxicity In Vitro.

N-methyl-d-aspartate receptors (NMDARs) are ionotropic glutamate receptors controlling fundamental physiological processes in the central nervous system, such as learning and memory. Excessive activation of NMDARs causes excitotoxicity and results in neurodegeneration, which is observed in a number of pathological conditions. Because of their dichotomous role, therapeutic targeting of NMDAR is difficult. However, several lines of evidence suggest that excitotoxicity is predominantly linked to extrasynaptically located NMDARs. Here, we report on a nanoparticle-based strategy to inhibit extrasynaptic NMDARs exclusively and subtype selectively, while allowing synaptic NMDARs activity. We designed gold nanoparticles (AuNPs) carrying conopeptide derivatives conjugated on their poly(ethylene glycol) coating as allosteric NMDAR inhibitors and show that these nanoparticles antagonize exclusively extrasynaptic NMDAR-mediated currents in cultured hippocampal neurons. Additionally, we show that conopeptide-functionalized AuNPs are neuroprotective in an in vitro model of excitotoxicity. By using AuNPs carrying different allosteric inhibitors with distinct NMDAR subtype selectivity such as peptide conantokin-G or peptide conantokin-R, we suggest activation of extrasynaptic GluN2B-containing diheteromeric NMDARs as the main cause of excitotoxicity.

Using well-defined Ag nanocubes as substrates to quantify the spatial resolution and penetration depth of surface-enhanced Raman scattering imaging.

The multiplexing capability and high sensitivity of surface-enhanced Raman scattering (SERS) make this new imaging modality particularly attractive for rapid diagnosis. With 100 nm Ag nanocubes serving as the substrate, this work quantitatively evaluated, for the first time, some of the fundamental parameters of SERS imaging such as blur, spatial resolution and penetration depth. Our results imply that SERS is a high-resolution imaging technique with a blur value of 0.5 µm that is lower than many traditional modalities such as mammography. The spatial resolution was measured to be 1.1 µm, suggesting that SERS images could be collected effectively by adjusting the imaging step size to the same length scale, or no more than 2 µm. The major drawback of SERS imaging is its penetration depth, which is limited by the scattering and absorption of tissues. We demonstrated that enhancement of signal caused by aggregation of multiple nanoparticles could help overcome this potential road-block to in vivo imaging.

Plasmonic near-electric field enhancement effects in ultrafast photoelectron emission: correlated spatial and laser polarization microscopy studies of individual Ag nanocubes.

Electron emission from single, supported Ag nanocubes excited with ultrafast laser pulses (λ = 800 nm) is studied via spatial and polarization correlated (i) dark field scattering microscopy (DFM), (ii) scanning photoionization microscopy (SPIM), and (iii) high-resolution transmission electron microscopy (HRTEM). Laser-induced electron emission is found to peak for laser polarization aligned with cube diagonals, suggesting the critical influence of plasmonic near-field enhancement of the incident electric field on the overall electron yield. For laser pulses with photon energy below the metal work function, coherent multiphoton photoelectron emission (MPPE) is identified as the most probable mechanism responsible for electron emission from Ag nanocubes and likely metal nanoparticles/surfaces in general.

Gold nanocages: from synthesis to theranostic applications.

Gold nanostructures have garnered considerable attention in recent years for their potential to facilitate both the diagnosis and treatment of cancer through their advantageous chemical and physical properties. The key feature of Au nanostructures for enabling this diverse array of biomedical applications is their attractive optical properties, specifically the scattering and absorption of light at resonant wavelengths due to the excitation of plasmon oscillations. This phenomenon is commonly known as localized surface plasmon resonance (LSPR) and is the source of the ruby red color of conventional Au colloids. The resonant wavelength depends on the size, shape, and geometry of the nanostructures, providing a set of knobs to manipulate the optical properties as needed. For in vivo applications, especially when optical excitation or transduction is involved, the LSPR peaks of the Au nanostructures have to be tuned to the transparent window of soft tissues in the near-infrared (NIR) region (from 700 to 900 nm) to maximize the penetration depth. Gold nanocages represent one class of nanostructures with tunable LSPR peaks in the NIR region. These versatile nanostructures, characterized by hollow interiors and ultrathin, porous walls, can be prepared in relatively large quantities using a remarkably simple procedure based on the galvanic replacement between Ag nanocubes and aqueous chloroauric acid. The LSPR peaks of Au nanocages can be readily and precisely tuned to any wavelength in the NIR region by controlling their size, wall thickness, or both. Other significant features of Au nanocages that make them particularly intriguing materials for biomedical applications include their compact sizes, large absorption cross sections (almost five orders of magnitude greater than those of conventional organic dyes), and their bio-inertness, as well as a robust and straightforward procedure for surface modification based on Au-thiolate chemistry. In this Account, we present some of the most recent advances in the use of Au nanocages for a broad range of theranostic applications. First, we describe their use as tracers for tracking by multiphoton luminescence. Gold nanocages can also serve as contrast agents for photoacoustic (PA) and mutimodal (PA/fluorescence) imaging. In addition, these nanostructures can be used as photothermal agents for the selective destruction of cancerous or diseased tissue. Finally, Au nanocages can serve as drug delivery vehicles for controlled and localized release in response to external stimuli such as NIR radiation or high-intensity focused ultrasound (HIFU).

The role of surface nonuniformity in controlling the initiation of a galvanic replacement reaction.

The use of silver nanocrystals--asymmetrically truncated octahedrons and nanobars--characterized by a nonuniform surface as substrates for a galvanic replacement reaction was investigated. As the surfaces of these nanocrystals contain facets with a variety of different areas, shapes, and atomic arrangements, we were able to examine the roles of these parameters in different stages of the galvanic replacement reaction with HAuCl(4) (e.g., pitting, hollowing, pit closing, and pore formation), and thus obtain a deeper understanding of the reaction mechanism than is possible with silver nanocubes. We found that the most important of these parameters was the atomic arrangement, that is, whether the surface was capped by a {100} or {111} facet, and that the area and shape of the facet had essentially no effect on the initiation of the reaction. Interestingly, through the reaction with asymmetrically truncated octahedrons, we were also able to demonstrate that even when pitting occurred over a large area, this region would be sealed through a combination of atomic diffusion and deposition during the intermediate stages of the reaction. Consequently, even if pitting occurred across a large percentage of the nanocrystal surface, it was still possible to maintain the morphology of the template throughout the reaction.

Gold nanostructures: a class of multifunctional materials for biomedical applications.

Gold nanostructures have proven to be a versatile platform for a broad range of biomedical applications, with potential use in numerous areas including: diagnostics and sensing, in vitro and in vivo imaging, and therapeutic techniques. These applications are possible because of the highly favorable properties of gold nanostructures, many of which can be tailored for specific applications. In the first part of this tutorial review, we will discuss the most critical properties of gold nanostructures for biomedical applications: surface chemistry, localized surface plasmon resonance (LSPR), and morphology. In the second part of the review, we will discuss how these properties can be harnessed for a selection of biomedical applications, aiming to give the reader an overview of general strategies as well as highlight some recent advances in this field.

Engineering the Properties of Metal Nanostructures via Galvanic Replacement Reactions.

In this review, we will bring the reader up to date with recent advances in the use of galvanic replacement reactions to engineer highly tunable nanostructures for a variety of applications. We will begin by discussing the variety of templates that have been used for such reactions and how the structural details (e.g., shape, size, and defects, among others) have interesting effects on the ultimate product, beyond serving as a simple site for deposition. This will be followed by a discussion of how we can manipulate the processes of alloying and dealloying to produce novel structures and how the type of precursor affects the final properties. Finally, the interesting optical properties of these materials and some innovative applications in areas of biomedical engineering and catalysis will be discussed, completing our overview of the state of the art in galvanic replacement.

In vivo molecular photoacoustic tomography of melanomas targeted by bioconjugated gold nanocages.

Early diagnosis, accurate staging, and image-guided resection of melanomas remain crucial clinical objectives for improving patient survival and treatment outcomes. Conventional techniques cannot meet this demand because of the low sensitivity, low specificity, poor spatial resolution, shallow penetration, and/or ionizing radiation. Here we overcome such limitations by combining high-resolution photoacoustic tomography (PAT) with extraordinarily optical absorbing gold nanocages (AuNCs). When bioconjugated with [Nle(4),D-Phe(7)]-alpha-melanocyte-stimulating hormone, the AuNCs can serve as a novel contrast agent for in vivo molecular PAT of melanomas with both exquisite sensitivity and high specificity. The bioconjugated AuNCs enhanced contrast approximately 300% more than the control, PEGylated AuNCs. The in vivo PAT quantification of the amount of AuNCs accumulated in melanomas was further validated with inductively coupled plasma mass spectrometry (ICP-MS).

Dissolving Ag from Au-Ag Alloy Nanoboxes with H(2)O(2): A Method for Both Tailoring the Optical Properties and Measuring the H(2)O(2) Concentration.

This article describes a method for generating Au-based nanocages with controlled wall thickness, porosity, and optical properties by dissolving Ag from Au-Ag alloy nanoboxes with H(2)O(2). It involves two steps: i) formation of Au-Ag alloy nanoboxes with some pure Ag left behind by titrating Ag nanocubes with aqueous HAuCl(4); and ii) removal of Ag atoms from both the pure Ag remaining in the nanoboxes and the alloy walls via H(2)O(2) etching. The optical properties of the resultant Au-Ag nanocages can be easily tailored by controlling the amount of H(2)O(2) added into the reaction system. Due to the changes to the optical spectra, the Au-Ag alloy nanoboxes can also be employed to detect H(2)O(2) with a more or less linear readout in the range of concentration from 5×10(-2) M down to 5×10(-7) M.

Targeting gold nanocages to cancer cells for photothermal destruction and drug delivery.

IMPORTANCE OF THE FIELD: Plasmonic nanoparticles provide a new route to treat cancer owing to their ability to convert light into heat effectively for photothermal destruction. Combined with the targeting mechanisms possible with nanoscale materials, this technique has the potential to enable highly targeted therapies to minimize undesirable side effects. AREAS COVERED IN THIS REVIEW: This review discusses the use of gold nanocages, a new class of plasmonic nanoparticles, for photothermal applications. Gold nanocages are hollow, porous structures with compact sizes and precisely controlled plasmonic properties and surface chemistry. Also, a recent study of gold nanocages as drug-release carriers by externally controlling the opening and closing of the pores with a smart polymer whose conformation changes at a specific temperature is discussed. Release of the contents can be initiated remotely through near-infrared irradiation. Together, these topics cover the years from 2002 to 2009. WHAT THE READER WILL GAIN: The reader will be exposed to different aspects of gold nanocages, including synthesis, surface modification, in vitro studies, initial in vivo data and perspectives on future studies. TAKE HOME MESSAGE: Gold nanocages are a promising platform for cancer therapy in terms of both photothermal destruction and drug delivery.

Gold Nanocages: A Novel Class of Multifunctional Nanomaterials for Theranostic Applications.

Gold nanocages represent a novel class of nanostructures, well-suited for biomedical applications. They can be readily prepared via the galvanic replacement reaction between silver nanocubes and chloroauric acid. Their optical resonance peaks can be easily and precisely tuned to the near-infrared region from 650-900 nm, the transparent window for blood and soft tissue. Furthermore, their surface can be conveniently conjugated with various ligands for targeting cancer. In this feature article, we highlight recent advances in the large-scale synthesis of gold nanocages and their applications in cancer diagnosis and treatment. Specifically, we have scaled up the production of gold nanocages for in vivo studies and evaluated their tumor targeting capabilities. We have also demonstrated their use as contrast agents for photoacoustic tumor imaging and the mapping of sentinel lymph node, as photothermal transducers for cancer treatment, and as smart carriers for controlled release with a near-infrared laser.

Quantifying the cellular uptake of antibody-conjugated Au nanocages by two-photon microscopy and inductively coupled plasma mass spectrometry.

Gold nanocages with localized surface plasmon resonance peaks in the near-infrared region exhibited a broad two-photon photoluminescence band extending from 450 to 650 nm when excited by a Ti:sapphire laser at 800 nm. The bright luminescence makes it possible to explore the use of Au nanocages as a new class of optical imaging agents for two-photon microscopy. In this work, we have demonstrated the use of two-photon microscopy as a convenient tool to directly examine the uptake of antibody-conjugated and PEGylated Au nanocages by U87MGwtEGFR cells. We have also correlated the results from two-photon microscopy with the data obtained by inductively coupled plasma mass spectrometry. Combined together, these results indicate that the antibody-conjugated Au nanocages were attached to the surface of the cells through antibody-antigen binding and then internalized into the cells via receptor-mediated endocytosis. The cellular uptake process was dependent on a number of parameters, including incubation time, incubation temperature, size of the Au nanocages, and the number of antibodies immobilized on each nanocage.

Gold nanocages covered by smart polymers for controlled release with near-infrared light.

Photosensitive caged compounds have enhanced our ability to address the complexity of biological systems by generating effectors with remarkable spatial/temporal resolutions. The caging effect is typically removed by photolysis with ultraviolet light to liberate the bioactive species. Although this technique has been successfully applied to many biological problems, it suffers from a number of intrinsic drawbacks. For example, it requires dedicated efforts to design and synthesize a precursor compound for each effector. The ultraviolet light may cause damage to biological samples and is suitable only for in vitro studies because of its quick attenuation in tissue. Here we address these issues by developing a platform based on the photothermal effect of gold nanocages. Gold nanocages represent a class of nanostructures with hollow interiors and porous walls. They can have strong absorption (for the photothermal effect) in the near-infrared while maintaining a compact size. When the surface of a gold nanocage is covered with a smart polymer, the pre-loaded effector can be released in a controllable fashion using a near-infrared laser. This system works well with various effectors without involving sophisticated syntheses, and is well suited for in vivo studies owing to the high transparency of soft tissue in the near-infrared region.

Measuring the surface-enhanced Raman scattering enhancement factors of hot spots formed between an individual Ag nanowire and a single Ag nanocube.

This paper describes a systematic study of the surface-enhanced Raman scattering (SERS) activity of hot spots formed between a Ag nanowire and a Ag nanocube with sharp corners. We investigated two distinct dimer structures: (i) a nanocube having one side face nearly touching the side face of a nanowire, and (ii) a nanocube having one edge nearly touching the side face of a nanowire. The field enhancements for the dimers displayed a strong dependence on laser polarization, and the strongest SERS intensities were observed for polarization along the hot-spot axis. Moreover, the detected SERS intensities were dependent on the hot-spot structure, i.e., the relative orientation of the Ag nanocube with respect to the nanowire's side face. When the dimer had a face-to-face configuration, the enhancement factor EF(dimer) was 1.4 x 10(7). This corresponds to 22-fold and 24-fold increases compared to those for individual Ag nanowires and nanocubes, respectively. Conversely, when the dimer had an edge-to-face configuration, EF(dimer) was 4.3 x 10(6). These results demonstrated that the number of probe molecules adsorbed at the hot spot played an important role in determining the detected SERS intensities. EF(dimer) was maximized when the dimer configuration allowed for a larger number of probe molecules to be trapped within the hot-spot region.