Remotely controlled drug release in deep brain regions of non-human primates.

Many areas of science and medicine would benefit from selective release of drugs in specific regions. Nanoparticle drug carriers activated by focused ultrasound-remotely applied, depth-penetrating energy-may provide such selective interventions. Here, we developed stable, ultrasound-responsive nanoparticles that can be used to release drugs effectively and safely in non-human primates. The nanoparticles were used to release propofol in deep brain visual regions. The release reversibly modulated the subjects' visual choice behavior and was specific to the targeted region and to the released drug. Gadolinium-enhanced MR imaging suggested an intact blood-brain barrier. Blood draws showed normal clinical chemistry and hematology. In summary, this study provides a safe and effective approach to release drugs on demand in selected deep brain regions at levels sufficient to modulate behavior.

Matching provides efficient decisions.

How humans and animals distribute their behavior across choice options has been of key interest to economics, psychology, ecology, and related fields. Neoclassical and behavioral economics have provided prescriptions for how decision-makers can maximize their reward or utility, but these formalisms are used by decision-makers rarely. Instead, individuals allocate their behavior in proportion to the worth of their options, a phenomenon captured by the generalized matching law. Why biological decision-makers adopt this strategy has been unclear. To provide insight into this issue, this article evaluates the performance of matching across a broad spectrum of decision situations, using simulations. Matching is found to attain a high or near-optimal gain, and the strategy achieves this level of performance following a single evaluation of the decision options. Thus, matching provides highly efficient decisions across a wide range of choice environments. This result offers a quantitative explanation for the broad adoption of matching by biological decision-makers.

Matching provides efficient decisions.

How humans and animals distribute their behavior across choice options has been of key interest to economics, psychology, ecology, and related fields. Neoclassical and behavioral economics have provided prescriptions for how decision-makers can maximize their reward or utility, but these formalisms are used by decision-makers rarely. Instead, individuals allocate their behavior in proportion to the worth of their options, a phenomenon captured by the generalized matching law. Why biological decision-makers adopt this strategy has been unclear. To provide insight into this issue, this article evaluates the performance of matching across a broad spectrum of decision situations, using simulations. Matching is found to attain a high or near-optimal gain, and the strategy achieves this level of performance following a single evaluation of the decision options. Thus, matching provides highly efficient decisions across a wide range of choice environments. This result offers a quantitative explanation for the broad adoption of matching by biological decision-makers.

Noninvasive modulation of essential tremor with focused ultrasonic waves.

Objective: Transcranial focused low-intensity ultrasound has the potential to noninvasively modulate confined regions deep inside the human brain, which could provide a new tool for causal interrogation of circuit function in humans. However, it has been unclear whether the approach is potent enough to modulate behavior.Approach: To test this, we applied low-intensity ultrasound to a deep brain thalamic target, the ventral intermediate nucleus, in three patients with essential tremor.Main results: Brief, 15 s stimulations of the target at 10% duty cycle with low-intensity ultrasound, repeated less than 30 times over a period of 90 min, nearly abolished tremor (98% and 97% tremor amplitude reduction) in 2 out of 3 patients. The effect was observed within seconds of the stimulation onset and increased with ultrasound exposure time. The effect gradually vanished following the stimulation, suggesting that the stimulation was safe with no harmful long-term consequences detected.Significance: This result demonstrates that low-intensity focused ultrasound can robustly modulate deep brain regions in humans with notable effects on overt motor behavior.

Device for Multifocal Delivery of Ultrasound Into Deep Brain Regions in Humans.

Low-intensity focused ultrasound provides the means to noninvasively stimulate or release drugs in specified deep brain targets. However, successful clinical translations require hardware that maximizes acoustic transmission through the skull, enables flexible electronic steering, and provides accurate and reproducible targeting while minimizing the use of MRI. We have developed a device that addresses these practical requirements. The device delivers ultrasound through the temporal and parietal skull windows, which minimize the attenuation and distortions of the ultrasound by the skull. The device consists of 252 independently controlled elements, which provides the ability to modulate multiple deep brain targets at a high spatiotemporal resolution, without the need to move the device or the subject. And finally, the device uses a mechanical registration method that enables accurate deep brain targeting both inside and outside of the MRI. Using this method, a single MRI scan is necessary for accurate targeting; repeated subsequent treatments can be performed reproducibly in an MRI-free manner. We validated these functions by transiently modulating specific deep brain regions in two patients with treatment-resistant depression.

Remotely controlled drug release in deep brain regions of non-human primates.

Many areas of science and medicine would benefit from selective release of drugs in specific regions of interest. Nanoparticle drug carriers activated by focused ultrasound-remotely applied, depth-penetrating energy-may provide such selective interventions. Here, we developed stable, ultrasound-responsive nanoparticles that can be used to release drugs effectively and safely in non-human primates. The nanoparticles were used to release propofol in deep brain visual regions. The release reversibly modulated the subjects' visual choice behavior and was specific to the targeted region and to the released drug. Gadolinium-enhanced MRI imaging suggested an intact blood-brain barrier. Blood draws showed normal clinical chemistry and hematology. In summary, this study provides a safe and effective approach to release drugs on demand in selected deep brain regions at levels sufficient to modulate behavior.

Durable effects of deep brain ultrasonic neuromodulation on major depression: a case report.

BACKGROUND: Severe forms of depression have been linked to hyperactivity of the subcallosal cingulate cortex. The ability to stimulate the subcallosal cingulate cortex or associated circuits noninvasively and directly would maximize the number of patients who could receive treatment. To this end, we have developed an ultrasound-based device for effective noninvasive modulation of deep brain circuits. Here we describe an application of this tool to an individual with treatment-resistant depression. CASE PRESENTATION: A 30-year-old Caucasian woman with severe treatment-resistant non-psychotic depression was recruited into a clinical study approved by the Institutional Review Board of the University of Utah. The patient had a history of electroconvulsive therapy with full remission but without sustained benefit. Magnetic resonance imaging was used to coregister the ultrasound device to the subject's brain anatomy and to evaluate neural responses to stimulation. Brief, 30-millisecond pulses of low-intensity ultrasound delivered into the subcallosal cingulate cortex target every 4 seconds caused a robust decrease in functional magnetic resonance imaging blood-oxygen-level-dependent activity within the target. Following repeated stimulation of three anterior cingulate targets, the patient's depressive symptoms resolved within 24 hours of the stimulation. The patient remained in remission for at least 44 days afterwards. CONCLUSIONS: This case illustrates the potential for ultrasonic neuromodulation to precisely engage deep neural circuits and to trigger a durable therapeutic reset of those circuits. Trial registration ClinicalTrials.gov, NCT05301036. Registered 29 March 2022, https://clinicaltrials.gov/ct2/show/NCT05301036.

Stem cell-derived brain organoids for controlled studies of transcranial neuromodulation.

Transcranial neuromodulation methods have the potential to diagnose and treat brain disorders at their neural source in a personalized manner. However, it has been difficult to investigate the direct effects of transcranial neuromodulation on neurons in human brain tissue. Here, we show that human brain organoids provide a detailed and artifact-free window into neuromodulation-evoked electrophysiological effects. We derived human cortical organoids from induced pluripotent stem cells and implanted 32-channel electrode arrays. Each organoid was positioned in the center of the human skull and subjected to low-intensity transcranial focused ultrasound. We found that ultrasonic stimuli modulated network activity in the gamma and delta ranges of the frequency spectrum. The effects on the neural networks were a function of the ultrasound stimulation frequency. High gamma activity remained elevated for at least 20 minutes following stimulation offset. This approach is expected to provide controlled studies of the effects of ultrasound and other transcranial neuromodulation modalities on human brain tissue.

Remote targeted electrical stimulation.

Objective:The ability to generate electric fields in specific targets remotely would transform manipulations of processes that rest on electrical signaling.Approach:This article shows that focal electric fields are generated from distance by combining two orthogonal, remotely applied energies-magnetic and focused ultrasonic fields. The effect derives from the Lorentz force equation applied to magnetic and ultrasonic fields.Main results:We elicited this effect using standard hardware and confirmed that the generated electric fields align with the Lorentz equation. The effect significantly and safely modulated human peripheral nerves and deep brain regions of non-human primates.Significance:This approach opens a new set of applications in which electric fields are generated at high spatiotemporal resolution within intact biological tissues or materials, thus circumventing the limitations of traditional electrode-based procedures.

Sustained modulation of primate deep brain circuits with focused ultrasonic waves.

BACKGROUND: Transcranial focused ultrasound has the potential to noninvasively modulate deep brain circuits and impart sustained, neuroplastic effects. OBJECTIVE: Bring the approach closer to translations by demonstrating sustained modulation of deep brain circuits and choice behavior in task-performing non-human primates. METHODS: Low-intensity transcranial ultrasound of 30 s in duration was delivered in a controlled manner into deep brain targets (left or right lateral geniculate nucleus; LGN) of non-human primates while the subjects decided whether a left or a right visual target appeared first. While the animals performed the task, we recorded intracranial EEG from occipital screws. The ultrasound was delivered into the deep brain targets daily for a period of more than 6 months. RESULTS: The brief stimulation induced effects on choice behavior that persisted up to 15 minutes and were specific to the sonicated target. Stimulation of the left/right LGN increased the proportion of rightward/leftward choices. These effects were accompanied by an increase in gamma activity over visual cortex. The contralateral effect on choice behavior and the increase in gamma, compared to sham stimulation, suggest that the stimulation excited the target neural circuits. There were no detrimental effects on the animals' discrimination performance over the months-long course of the stimulation. CONCLUSION: This study demonstrates that brief, 30-s ultrasonic stimulation induces neuroplastic effects specifically in the target deep brain circuits, and that the stimulation can be applied daily without detrimental effects. These findings encourage repeated applications of transcranial ultrasound to malfunctioning deep brain circuits in humans with the goal of providing a durable therapeutic reset.

Multifrequency-based sharpening of focal volume.

Systems that emit electromagnetic or sonic waves for diagnostic or interventional applications often have constraints on the size of their aperture, and thus produce an elongated focus in the axial dimension. This extended depth of focus limits imaging resolution and spatial specificity of the delivered energy. Here, we have developed a method that substantially minimizes the depth of focus. The method superimposes beams of distinct frequencies in space and time to create constructive interference at target and amplify deconstructive interference everywhere else, thus sharpening the focus. The method does not require labeling of targets or other manipulations of the medium. Using simulations, we found that the method tightens the depth of focus even for systems with a narrow bandwidth. Moreover, we implemented the method in ultrasonic hardware and found that a 46.1% frequency fractional bandwidth provides an average 7.4-fold reduction in the focal volume of the resulting beams. This method can be readily applied to sharpen the focus of interventional systems and is expected to also improve the axial resolution of existing imaging systems.

Transcranial ultrasound neuromodulation of the thalamic visual pathway in a large animal model and the dose-response relationship with MR-ARFI.

Neuromodulation of deep brain structures via transcranial ultrasound stimulation (TUS) is a promising, but still elusive approach to non-invasive treatment of brain disorders. The purpose of this study was to confirm that MR-guided TUS of the lateral geniculate nucleus (LGN) can modulate visual evoked potentials (VEPs) in the intact large animal; and to study the impact on cortical brain oscillations. The LGN on one side was identified with T2-weighted MRI in sheep (all male, n = 9). MR acoustic radiation force imaging (MR-ARFI) was used to confirm localization of the targeted area in the brain. Electroencephalographic (EEG) signals were recorded, and the visual evoked potential (VEP) peak-to-peak amplitude (N70 and P100) was calculated for each trial. Time-frequency spectral analysis was performed to elucidate the effect of TUS on cortical brain dynamics. The VEP peak-to-peak amplitude was reversibly suppressed relative to baseline during TUS. Dynamic spectral analysis demonstrated a change in cortical oscillations when TUS is paired with visual sensory input. Sonication-associated microscopic displacements, as measured by MR-ARFI, correlated with the TUS-mediated suppression of visual evoked activity. TUS non-invasively delivered to LGN can neuromodulate visual activity and oscillatory dynamics in large mammalian brains.

Remus: System for remote deep brain interventions.

Transcranial-focused ultrasound brings personalized medicine to the human brain. Ultrasound can modulate neural activity or release drugs in specific neural circuits but this personalized approach requires a system that delivers ultrasound into specified targets flexibly and on command. We developed a remote ultrasound system (Remus) that programmatically targets deep brain regions with high spatiotemporal precision and in a multi-focal manner. We validated these functions by modulating two deep brain nuclei-the left and right lateral geniculate nucleus-in a task-performing nonhuman primate. This flexible system will enable researchers and clinicians to diagnose and treat specific deep brain circuits in a noninvasive yet targeted manner, thus embodying the promise of personalized treatments of brain disorders.

Modeling human telencephalic development and autism-associated SHANK3 deficiency using organoids generated from single neural rosettes.

Human telencephalon is an evolutionarily advanced brain structure associated with many uniquely human behaviors and disorders. However, cell lineages and molecular pathways implicated in human telencephalic development remain largely unknown. We produce human telencephalic organoids from stem cell-derived single neural rosettes and investigate telencephalic development under normal and pathological conditions. We show that single neural rosette-derived organoids contain pallial and subpallial neural progenitors, excitatory and inhibitory neurons, as well as macroglial and periendothelial cells, and exhibit predictable organization and cytoarchitecture. We comprehensively characterize the properties of neurons in SNR-derived organoids and identify transcriptional programs associated with the specification of excitatory and inhibitory neural lineages from a common pool of NPs early in telencephalic development. We also demonstrate that neurons in organoids with a hemizygous deletion of an autism- and intellectual disability-associated gene SHANK3 exhibit intrinsic and excitatory synaptic deficits and impaired expression of several clustered protocadherins. Collectively, this study validates SNR-derived organoids as a reliable model for studying human telencephalic cortico-striatal development and identifies intrinsic, synaptic, and clustered protocadherin expression deficits in human telencephalic tissue with SHANK3 hemizygosity.

Acoustic properties across the human skull.

Transcranial ultrasound is emerging as a noninvasive tool for targeted treatments of brain disorders. Transcranial ultrasound has been used for remotely mediated surgeries, transient opening of the blood-brain barrier, local drug delivery, and neuromodulation. However, all applications have been limited by the severe attenuation and phase distortion of ultrasound by the skull. Here, we characterized the dependence of the aberrations on specific anatomical segments of the skull. In particular, we measured ultrasound propagation properties throughout the perimeter of intact human skulls at 500 kHz. We found that the parietal bone provides substantially higher transmission (average pressure transmission 31 ± 7%) and smaller phase distortion (242 ± 44 degrees) than frontal (13 ± 2%, 425 ± 47 degrees) and occipital bone regions (16 ± 4%, 416 ± 35 degrees). In addition, we found that across skull regions, transmission strongly anti-correlated (R=-0.79) and phase distortion correlated (R=0.85) with skull thickness. This information guides the design, positioning, and skull correction functionality of next-generation devices for effective, safe, and reproducible transcranial focused ultrasound therapies.

Effective Ultrasonic Stimulation in Human Peripheral Nervous System.

OBJECTIVE: Low-intensity ultrasound can stimulate excitable cells in a noninvasive and targeted manner, but which parameters are effective has remained elusive. This question has been difficult to answer because differences in transducers and parameters-frequency in particular-lead to profound differences in the stimulated tissue volumes. The objective of this study is to control for these differences and evaluate which ultrasound parameters are effective in stimulating excitable cells. METHODS: Here, we stimulated the human peripheral nervous system using a single transducer operating in a range of frequencies, and matched the stimulated volumes with an acoustic aperture. RESULTS: We found that low frequencies (300 kHz) are substantially more effective in generating tactile and nociceptive responses in humans compared to high frequencies (900 kHz). The strong effect of ultrasound frequency was observed for all pressures tested, for continuous and pulsed stimuli, and for tactile and nociceptive responses. CONCLUSION: This prominent effect may be explained by a mechanical force associated with ultrasound. The effect is not due to heating, which would be weaker at the low frequency. SIGNIFICANCE: This controlled study reveals that ultrasonic stimulation of excitable cells is stronger at lower frequencies, which guides the choice of transducer hardware for effective ultrasonic stimulation of the peripheral nervous system in humans.

Remote, brain region-specific control of choice behavior with ultrasonic waves.

The ability to modulate neural activity in specific brain circuits remotely and systematically could revolutionize studies of brain function and treatments of brain disorders. Sound waves of high frequencies (ultrasound) have shown promise in this respect, combining the ability to modulate neuronal activity with sharp spatial focus. Here, we show that the approach can have potent effects on choice behavior. Brief, low-intensity ultrasound pulses delivered noninvasively into specific brain regions of macaque monkeys influenced their decisions regarding which target to choose. The effects were substantial, leading to around a 2:1 bias in choices compared to the default balanced proportion. The effect presence and polarity was controlled by the specific target region. These results represent a critical step towards the ability to influence choice behavior noninvasively, enabling systematic investigations and treatments of brain circuits underlying disorders of choice.

Histologic safety of transcranial focused ultrasound neuromodulation and magnetic resonance acoustic radiation force imaging in rhesus macaques and sheep.

BACKGROUND: Neuromodulation by transcranial focused ultrasound (FUS) offers the potential to non-invasively treat specific brain regions, with treatment location verified by magnetic resonance acoustic radiation force imaging (MR-ARFI). OBJECTIVE: To investigate the safety of these methods prior to widespread clinical use, we report histologic findings in two large animal models following FUS neuromodulation and MR-ARFI. METHODS: Two rhesus macaques and thirteen Dorset sheep were studied. FUS neuromodulation was targeted to the primary visual cortex in rhesus macaques and to subcortical locations, verified by MR-ARFI, in eleven sheep. Both rhesus macaques and five sheep received a single FUS session, whereas six sheep received repeated sessions three to six days apart. The remaining two control sheep did not receive ultrasound but otherwise underwent the same anesthetic and MRI procedures as the eleven experimental sheep. Hematoxylin and eosin-stained sections of brain tissue (harvested zero to eleven days following FUS) were evaluated for tissue damage at FUS and control locations as well as tissue within the path of the FUS beam. TUNEL staining was used to evaluate for the presence of apoptosis in sheep receiving high dose FUS. RESULTS: No FUS-related pre-mortem histologic findings were observed in the rhesus macaques or in any of the examined sheep. Extravascular red blood cells (RBCs) were present within the meninges of all sheep, regardless of treatment group. Similarly, small aggregates of perivascular RBCs were rarely noted in non-target regions of neural parenchyma of FUS-treated (8/11) and untreated (2/2) sheep. However, no concurrent histologic abnormalities were observed, consistent with RBC extravasation occurring as post-mortem artifact following brain extraction. Sheep within the high dose FUS group were TUNEL-negative at the targeted site of FUS. CONCLUSIONS: The absence of FUS-related histologic findings suggests that the neuromodulation and MR-ARFI protocols evaluated do not cause tissue damage.

Neuromodulation with transcranial focused ultrasound.

The understanding of brain function and the capacity to treat neurological and psychiatric disorders rest on the ability to intervene in neuronal activity in specific brain circuits. Current methods of neuromodulation incur a tradeoff between spatial focus and the level of invasiveness. Transcranial focused ultrasound (FUS) is emerging as a neuromodulation approach that combines noninvasiveness with focus that can be relatively sharp even in regions deep in the brain. This may enable studies of the causal role of specific brain regions in specific behaviors and behavioral disorders. In addition to causal brain mapping, the spatial focus of FUS opens new avenues for treatments of neurological and psychiatric conditions. This review introduces existing and emerging FUS applications in neuromodulation, discusses the mechanisms of FUS effects on cellular excitability, considers the effects of specific stimulation parameters, and lays out the directions for future work.

Ultrasound Elicits Behavioral Responses through Mechanical Effects on Neurons and Ion Channels in a Simple Nervous System.

Focused ultrasound has been shown to stimulate excitable cells, but the biophysical mechanisms behind this phenomenon remain poorly understood. To provide additional insight, we devised a behavioral-genetic assay applied to the well-characterized nervous system of Caenorhabditis elegans nematodes. We found that pulsed ultrasound elicits robust reversal behavior in wild-type animals in a pressure-, duration-, and pulse protocol-dependent manner. Responses were preserved in mutants unable to sense thermal fluctuations and absent in mutants lacking neurons required for mechanosensation. Additionally, we found that the worm's response to ultrasound pulses rests on the expression of MEC-4, a DEG/ENaC/ASIC ion channel required for touch sensation. Consistent with prior studies of MEC-4-dependent currents in vivo, the worm's response was optimal for pulses repeated 300-1000 times per second. Based on these findings, we conclude that mechanical, rather than thermal, stimulation accounts for behavioral responses. Further, we propose that acoustic radiation force governs the response to ultrasound in a manner that depends on the touch receptor neurons and MEC-4-dependent ion channels. Our findings illuminate a complete pathway of ultrasound action, from the forces generated by propagating ultrasound to an activation of a specific ion channel. The findings further highlight the importance of optimizing ultrasound pulsing protocols when stimulating neurons via ion channels with mechanosensitive properties.SIGNIFICANCE STATEMENT How ultrasound influences neurons and other excitable cells has remained a mystery for decades. Although it is widely understood that ultrasound can heat tissues and induce mechanical strain, whether or not neuronal activation depends on heat, mechanical force, or both physical factors is not known. We harnessed Caenorhabditis elegans nematodes and their extraordinary sensitivity to thermal and mechanical stimuli to address this question. Whereas thermosensory mutants respond to ultrasound similar to wild-type animals, mechanosensory mutants were insensitive to ultrasound stimulation. Additionally, stimulus parameters that accentuate mechanical effects were more effective than those producing more heat. These findings highlight a mechanical nature of the effect of ultrasound on neurons and suggest specific ways to optimize stimulation protocols in specific tissues.