No abstract

Ultrasound has received widespread attention as an emerging technology for targeted, non-invasive neuromodulation based on its ability to evoke electrophysiological and motor responses in animals. However, little is known about the spatiotemporal pattern of ultrasound-induced brain activity that could drive these responses. Here, we address this question by combining focused ultrasound with wide-field optical imaging of calcium signals in transgenic mice. Surprisingly, we find cortical activity patterns consistent with indirect activation of auditory pathways rather than direct neuromodulation at the ultrasound focus. Ultrasound-induced activity is similar to that evoked by audible sound. Furthermore, both ultrasound and audible sound elicit motor responses consistent with a startle reflex, with both responses reduced by chemical deafening. These findings reveal an indirect auditory mechanism for ultrasound-induced cortical activity and movement requiring careful consideration in future development of ultrasonic neuromodulation as a tool in neuroscience research.

Ultrasound is among the most widely used biomedical imaging modalities, but has limited ability to image specific molecular targets due to the lack of suitable nanoscale contrast agents. Gas vesicles – genetically encoded protein nanostructures isolated from buoyant photosynthetic microbes – have recently been identified as nanoscale reporters for ultrasound. Their unique physical properties give gas vesicles significant advantages over conventional microbubble contrast agents, including nanoscale dimensions and inherent physical stability. Furthermore, as a genetically encoded material, gas vesicles present the possibility that the nanoscale mechanical, acoustic and targeting properties of an imaging agent can be engineered at the level of its constituent proteins. Here, we demonstrate that genetic engineering of gas vesicles results in nanostructures with new mechanical, acoustic, surface and functional properties to enable harmonic, multiplexed and multimodal ultrasound imaging, as well as cell-specific molecular targeting. These results establish a biomolecular platform for the engineering of acoustic nanomaterials.

No abstract

Gas vesicles are a unique class of gas-filled protein nanostructures whose physical properties allow them to serve as highly sensitive imaging agents for ultrasound and magnetic resonance imaging (MRI), detectable at sub-nanomolar concentrations. Here we provide a protocol for isolating gas vesicles from native and heterologous host organisms, functionalizing these nanostructures with moieties for targeting and fluorescence, characterizing their biophysical properties and imaging them using ultrasound and magnetic resonance imaging. Gas vesicles can be isolated from natural cyanobacterial and haloarchaeal host organisms or from E. coli expressing a heterologous gas vesicle gene cluster, and purified using buoyancy-assisted techniques. They can then be modified by replacing surface-bound proteins with engineered, heterologously expressed variants, or through chemical conjugation, resulting in altered mechanical, surface and targeting properties. Pressurized absorbance spectroscopy is used to characterize their mechanical properties, while dynamic light scattering and transmission electron microscopy are used to determine nanoparticle size and morphology, respectively. Gas vesicles can then be imaged with ultrasound in vitro and in vivo using pulse sequences optimized for their detection versus background. They can also be imaged with hyperpolarized xenon MRI using chemical exchange saturation transfer between gas vesicle-bound and dissolved xenon – a technique currently implemented in vitro. Taking 3–8 days to prepare, these genetically encodable nanostructures enable multi-modal, noninvasive biological imaging with high sensitivity and potential for molecular targeting.

Nuclear spin polarization can be significantly increased through the process of hyperpolarization, leading to an increase in the sensitivity of nuclear magnetic resonance (NMR) experiments by 4–8 orders of magnitude. Hyperpolarized gases, unlike liquids and solids, can be more readily separated and purified from the compounds used to mediate the hyperpolarization processes. These pure hyperpolarized gases enabled many novel MRI applications including the visualization of void spaces, imaging of lung function, and remote detection. Additionally, hyperpolarized gases can be dissolved in liquids and can be used as sensitive molecular probes and reporters. This mini-review covers the fundamentals of the preparation of hyperpolarized gases and focuses on selected applications of interest to biomedicine and materials science.

Visualizing and modulating molecular and cellular processes occurring deep within living organisms is fundamental to our study of basic biology and disease. Currently, the most sophisticated tools available to dynamically monitor and control cellular events rely on light-responsive proteins, which are difficult to use outside of optically transparent model systems, cultured cells, or surgically accessed regions owing to strong scattering of light by biological tissue. In contrast, ultrasound is a widely used medical imaging and therapeutic modality that enables the observation and perturbation of internal anatomy and physiology but has historically had limited ability to monitor and control specific cellular processes. Recent advances are beginning to address this limitation through the development of biomolecular tools that allow ultrasound to connect directly to cellular functions such as gene expression. Driven by the discovery and engineering of new contrast agents, reporter genes, and bioswitches, the nascent field of biomolecular ultrasound carries a wave of exciting opportunities.

No abstract