Temporal interference stimulation disrupts spike timing in the primate brain.

Electrical stimulation can regulate brain activity, producing clear clinical benefits, but focal and effective neuromodulation often requires surgically implanted electrodes. Recent studies argue that temporal interference (TI) stimulation may provide similar outcomes non-invasively. During TI, scalp electrodes generate multiple electrical fields in the brain, modulating neural activity only at their intersection. Despite considerable enthusiasm for this approach, little empirical evidence demonstrates its effectiveness, especially under conditions suitable for human use. Here, using single-neuron recordings in non-human primates, we establish that TI reliably alters the timing, but not the rate, of spiking activity. However, we show that TI requires strategies-high carrier frequencies, multiple electrodes, and amplitude-modulated waveforms-that also limit its effectiveness. Combined, these factors make TI 80 % weaker than other forms of non-invasive brain stimulation. Although unlikely to cause widespread neuronal entrainment, TI may be ideal for disrupting pathological oscillatory activity, a hallmark of many neurological disorders.

The mode is the message.

In this issue of Neuron, Khazali et al.1 record neural activity during coordinated reaches and saccades. They find that excitatory neurons link arm and eye movement regions of parietal cortex, creating a multiregional mode that predicts movement timing and direction.

Transcranial electrical stimulation: How can a simple conductor orchestrate complex brain activity?

Transcranial electrical stimulation (tES) is one of the oldest and yet least understood forms of brain stimulation. The idea that a weak electrical stimulus, applied outside the head, can meaningfully affect neural activity is often regarded as mysterious. Here, we argue that the direct effects of tES are not so mysterious: Extensive data from a wide range of model systems shows it has appreciable effects on the activity of individual neurons. Instead, the real mysteries are how tES interacts with the brain's own activity and how these dynamics can be controlled to produce desirable therapeutic effects. These are challenging problems, akin to repairing a complex machine while it is running, but they are not unique to tES or even neuroscience. We suggest that models of coupled oscillators, a common tool for studying interactions in other fields, may provide valuable insights. By combining these tools with our growing, interdisciplinary knowledge of brain dynamics, we are now in a good position to make progress in this area and meet the high demand for effective neuromodulation in neuroscience and psychiatry.

Long-range cortical synchronization supports abrupt visual learning.

Visual plasticity declines sharply after the critical period, yet we easily learn to recognize new faces and places, even as adults. Such learning is often characterized by a "moment of insight," an abrupt and dramatic improvement in recognition. The mechanisms that support abrupt learning are unknown, but one hypothesis is that they involve changes in synchronization between brain regions. To test this hypothesis, we used a behavioral task in which non-human primates rapidly learned to recognize novel images and to associate them with specific responses. Simultaneous recordings from inferotemporal and prefrontal cortices revealed a transient synchronization of neural activity between these areas that peaked around the moment of insight. Synchronization was strongest between inferotemporal sites that encoded images and reward-sensitive prefrontal sites. Moreover, its magnitude intensified gradually over image exposures, suggesting that abrupt learning is the culmination of a search for informative signals within a circuit linking sensory information to task demands.

Brain stimulation competes with ongoing oscillations for control of spike timing in the primate brain.

Transcranial alternating current stimulation (tACS) is a popular method for modulating brain activity noninvasively. In particular, tACS is often used as a targeted intervention that enhances a neural oscillation at a specific frequency to affect a particular behavior. However, these interventions often yield highly variable results. Here, we provide a potential explanation for this variability: tACS competes with the brain's ongoing oscillations. Using neural recordings from alert nonhuman primates, we find that when neural firing is independent of ongoing brain oscillations, tACS readily entrains spiking activity, but when neurons are strongly entrained to ongoing oscillations, tACS often causes a decrease in entrainment instead. Consequently, tACS can yield categorically different results on neural activity, even when the stimulation protocol is fixed. Mathematical analysis suggests that this competition is likely to occur under many experimental conditions. Attempting to impose an external rhythm on the brain may therefore often yield precisely the opposite effect.

Neural Selectivity for Visual Motion in Macaque Area V3A.

The processing of visual motion is conducted by dedicated pathways in the primate brain. These pathways originate with populations of direction-selective neurons in the primary visual cortex, which projects to dorsal structures like the middle temporal (MT) and medial superior temporal (MST) areas. Anatomical and imaging studies have suggested that area V3A might also be specialized for motion processing, but there have been very few studies of single-neuron direction selectivity in this area. We have therefore performed electrophysiological recordings from V3A neurons in two macaque monkeys (one male and one female) and measured responses to a large battery of motion stimuli that includes translation motion, as well as more complex optic flow patterns. For comparison, we simultaneously recorded the responses of MT neurons to the same stimuli. Surprisingly, we find that overall levels of direction selectivity are similar in V3A and MT and moreover that the population of V3A neurons exhibits somewhat greater selectivity for optic flow patterns. These results suggest that V3A should be considered as part of the motion processing machinery of the visual cortex, in both human and non-human primates.

tACS entrains neural activity while somatosensory input is blocked.

Transcranial alternating current stimulation (tACS) modulates brain activity by passing electrical current through electrodes that are attached to the scalp. Because it is safe and noninvasive, tACS holds great promise as a tool for basic research and clinical treatment. However, little is known about how tACS ultimately influences neural activity. One hypothesis is that tACS affects neural responses directly, by producing electrical fields that interact with the brain's endogenous electrical activity. By controlling the shape and location of these electric fields, one could target brain regions associated with particular behaviors or symptoms. However, an alternative hypothesis is that tACS affects neural activity indirectly, via peripheral sensory afferents. In particular, it has often been hypothesized that tACS acts on sensory fibers in the skin, which in turn provide rhythmic input to central neurons. In this case, there would be little possibility of targeted brain stimulation, as the regions modulated by tACS would depend entirely on the somatosensory pathways originating in the skin around the stimulating electrodes. Here, we directly test these competing hypotheses by recording single-unit activity in the hippocampus and visual cortex of alert monkeys receiving tACS. We find that tACS entrains neuronal activity in both regions, so that cells fire synchronously with the stimulation. Blocking somatosensory input with a topical anesthetic does not significantly alter these neural entrainment effects. These data are therefore consistent with the direct stimulation hypothesis and suggest that peripheral somatosensory stimulation is not required for tACS to entrain neurons.

Transcranial alternating current stimulation entrains single-neuron activity in the primate brain.

Spike timing is thought to play a critical role in neural computation and communication. Methods for adjusting spike timing are therefore of great interest to researchers and clinicians alike. Transcranial electrical stimulation (tES) is a noninvasive technique that uses weak electric fields to manipulate brain activity. Early results have suggested that this technique can improve subjects' behavioral performance on a wide range of tasks and ameliorate some clinical conditions. Nevertheless, considerable skepticism remains about its efficacy, especially because the electric fields reaching the brain during tES are small, whereas the likelihood of indirect effects is large. Our understanding of its effects in humans is largely based on extrapolations from simple model systems and indirect measures of neural activity. As a result, fundamental questions remain about whether and how tES can influence neuronal activity in the human brain. Here, we demonstrate that tES, as typically applied to humans, affects the firing patterns of individual neurons in alert nonhuman primates, which are the best available animal model for the human brain. Specifically, tES consistently influences the timing, but not the rate, of spiking activity within the targeted brain region. Such effects are frequency- and location-specific and can reach deep brain structures; control experiments show that they cannot be explained by sensory stimulation or other indirect influences. These data thus provide a strong mechanistic rationale for the use of tES in humans and will help guide the development of future tES applications.

Immediate neurophysiological effects of transcranial electrical stimulation.

Noninvasive brain stimulation techniques are used in experimental and clinical fields for their potential effects on brain network dynamics and behavior. Transcranial electrical stimulation (TES), including transcranial direct current stimulation (tDCS) and transcranial alternating current stimulation (tACS), has gained popularity because of its convenience and potential as a chronic therapy. However, a mechanistic understanding of TES has lagged behind its widespread adoption. Here, we review data and modelling on the immediate neurophysiological effects of TES in vitro as well as in vivo in both humans and other animals. While it remains unclear how typical TES protocols affect neural activity, we propose that validated models of current flow should inform study design and artifacts should be carefully excluded during signal recording and analysis. Potential indirect effects of TES (e.g., peripheral stimulation) should be investigated in more detail and further explored in experimental designs. We also consider how novel technologies may stimulate the next generation of TES experiments and devices, thus enhancing validity, specificity, and reproducibility.

Transcranial Direct Current Stimulation Facilitates Associative Learning and Alters Functional Connectivity in the Primate Brain.

There has been growing interest in transcranial direct current stimulation (tDCS), a non-invasive technique purported to modulate neural activity via weak, externally applied electric fields. Although some promising preliminary data have been reported for applications ranging from stroke rehabilitation to cognitive enhancement, little is known about how tDCS affects the human brain, and some studies have concluded that it may have no effect at all. Here, we describe a macaque model of tDCS that allows us to simultaneously examine the effects of tDCS on brain activity and behavior. We find that applying tDCS to right prefrontal cortex improves monkeys' performance on an associative learning task. While firing rates do not change within the targeted area, tDCS does induce large low-frequency oscillations in the underlying tissue. These oscillations alter functional connectivity, both locally and between distant brain areas, and these long-range changes correlate with tDCS's effects on behavior. Together, these results are consistent with the idea that tDCS leads to widespread changes in brain activity and suggest that it may be a valuable method for cheaply and non-invasively altering functional connectivity in humans.

Single-cell profiling reveals that eRNA accumulation at enhancer-promoter loops is not required to sustain transcription.

Enhancers are intergenic DNA elements that regulate the transcription of target genes in response to signaling pathways by interacting with promoters over large genomic distances. Recent studies have revealed that enhancers are bi-directionally transcribed into enhancer RNAs (eRNAs). Using single-molecule fluorescence in situ hybridization (smFISH), we investigated the eRNA-mediated regulation of transcription during estrogen induction in MCF-7 cells. We demonstrate that eRNAs are localized exclusively in the nucleus and are induced with similar kinetics as target mRNAs. However, eRNAs are mostly nascent at enhancers and their steady-state levels remain lower than those of their cognate mRNAs. Surprisingly, at the single-allele level, eRNAs are rarely co-expressed with their target loci, demonstrating that active gene transcription does not require the continuous transcription of eRNAs or their accumulation at enhancers. When co-expressed, sub-diffraction distance measurements between nascent mRNA and eRNA signals reveal that co-transcription of eRNAs and mRNAs rarely occurs within closed enhancer-promoter loops. Lastly, basal eRNA transcription at enhancers, but not E2-induced transcription, is maintained upon depletion of MLL1 and ERα, suggesting some degree of chromatin accessibility prior to signal-dependent activation of transcription. Together, our findings suggest that eRNA accumulation at enhancer-promoter loops is not required to sustain target gene transcription.

Orexin Neurons Respond Differentially to Auditory Cues Associated with Appetitive versus Aversive Outcomes.

Orexin (Orx) neurons are known to be involved in the promotion and maintenance of waking because they discharge in association with cortical activation and muscle tone during waking and because, in their absence, waking with muscle tone cannot be maintained and narcolepsy with cataplexy ensues. Whether Orx neurons discharge during waking in association with particular conditions, notably with appetitive versus aversive stimuli or positive versus negative emotions, is debated and considered important in understanding their role in supporting particular waking behaviors. Here, we used the technique of juxtacellular recording and labeling in head-fixed rats to characterize the discharge of Orx neurons during the performance of an associative discrimination task with auditory cues for appetitive versus aversive outcomes. Of 57 active, recorded, and neurobiotin-labeled neurons in the lateral hypothalamus, 11 were immunohistochemically identified as Orx-positive (Orx(+)), whereas none were identified as melanin-concentrating hormone-positive. Orx(+) neurons discharged at significantly higher rates during the tone associated with sucrose than during the tone associated with quinine delivered upon licking. They also discharged at high rates after the tone associated with sucrose. Across periods and outcomes, their discharge was positively correlated with EEG gamma activity and EMG activity, which is indicative of cortical activation and behavioral arousal. These results suggest that Orx neurons discharge in a manner characteristic of reward neurons yet also characteristic of arousal neurons. Accordingly, the Orx neurons may respond to and participate in reward processes while modulating cortical activity and muscle tone to promote and maintain arousal along with learned adaptive behavioral responses. SIGNIFICANCE STATEMENT: Orexin neurons play a critical role in promoting and maintaining a waking state because, in their absence, narcolepsy with cataplexy ensues. Known to discharge during waking and not during sleep, they have also been proposed to be selectively active during appetitive behaviors. Here, we recorded and labeled neurons in rats to determine the discharge of immunohistochemically identified orexin neurons during performance of an associative discrimination task. Orexin neurons responded differentially to auditory cues associated with appetitive sucrose versus aversive quinine, indicating that they behave like reward neurons. However, correlated discharge with cortical and muscle activity indicates that they also behave like arousal neurons and can thus promote cortical activation with behavioral arousal and muscle tone during adaptive waking behaviors.

On comparing in vivo intracranial recordings in non-human primates to predictions of optimized transcranial electrical stimulation.

Transcranial electrical stimulation (tES) can be optimized to achieve maximal current flow at desired brain regions. The aim of this study was to characterize electric field magnitudes generated by tES optimization and to compare them to experimentally induced values as determined by data from intracranial electrodes. Local field potentials were recorded from two monkeys with implanted multi-site intracranial Utah arrays during transcranial direct current stimulation (tDCS), and the neural effect predictions obtained from optimized electrode placement were assessed. Comparative data between the two sites of intracranial recordings during tDCS partially validated the predictions of our tES optimization algorithms.

The nuclear basket mediates perinuclear mRNA scanning in budding yeast.

After synthesis and transit through the nucleus, messenger RNAs (mRNAs) are exported to the cytoplasm through the nuclear pore complex (NPC). At the NPC, messenger ribonucleoproteins (mRNPs) first encounter the nuclear basket where mRNP rearrangements are thought to allow access to the transport channel. Here, we use single mRNA resolution live cell microscopy and subdiffraction particle tracking to follow individual mRNAs on their path toward the cytoplasm. We show that when reaching the nuclear periphery, RNAs are not immediately exported but scan along the nuclear periphery, likely to find a nuclear pore allowing export. Deletion or mutation of the nuclear basket proteins MLP1/2 or the mRNA binding protein Nab2 changes the scanning behavior of mRNPs at the nuclear periphery, shortens residency time at nuclear pores, and results in frequent release of mRNAs back into the nucleoplasm. These observations suggest a role for the nuclear basket in providing an interaction platform that keeps RNAs at the periphery, possibly to allow mRNP rearrangements before export.

Myosin VI regulates gene pairing and transcriptional pause release in T cells.

Naive CD4 T cells differentiate into several effector lineages, which generate a stronger and more rapid response to previously encountered immunological challenges. Although effector function is a key feature of adaptive immunity, the molecular basis of this process is poorly understood. Here, we investigated the spatiotemporal regulation of cytokine gene expression in resting and restimulated effector T helper 1 (Th1) cells. We found that the Lymphotoxin (LT)/TNF alleles, which encode TNF-α, were closely juxtaposed shortly after T-cell receptor (TCR) engagement, when transcription factors are limiting. Allelic pairing required a nuclear myosin, myosin VI, which is rapidly recruited to the LT/TNF locus upon restimulation. Furthermore, transcription was paused at the TNF locus and other related genes in resting Th1 cells and released in a myosin VI-dependent manner following activation. We propose that homologous pairing and myosin VI-mediated transcriptional pause release account for the rapid and efficient expression of genes induced by an external stimulus.

Contextual modulation and stimulus selectivity in extrastriate cortex.

Contextual modulation is observed throughout the visual system, using techniques ranging from single-neuron recordings to behavioral experiments. Its role in generating feature selectivity within the retina and primary visual cortex has been extensively described in the literature. Here, we describe how similar computations can also elaborate feature selectivity in the extrastriate areas of both the dorsal and ventral streams of the primate visual system. We discuss recent work that makes use of normalization models to test specific roles for contextual modulation in visual cortex function. We suggest that contextual modulation renders neuronal populations more selective for naturalistic stimuli. Specifically, we discuss contextual modulation's role in processing optic flow in areas MT and MST and for representing naturally occurring curvature and contours in areas V4 and IT. We also describe how the circuitry that supports contextual modulation is robust to variations in overall input levels. Finally, we describe how this theory relates to other hypothesized roles for contextual modulation.

Surround suppression and sparse coding in visual and barrel cortices.

During natural vision the entire retina is stimulated. Likewise, during natural tactile behaviors, spatially extensive regions of the somatosensory surface are co-activated. The large spatial extent of naturalistic stimulation means that surround suppression, a phenomenon whose neural mechanisms remain a matter of debate, must arise during natural behavior. To identify common neural motifs that might instantiate surround suppression across modalities, we review models of surround suppression and compare the evidence supporting the competing ideas that surround suppression has either cortical or sub-cortical origins in visual and barrel cortex. In the visual system there is general agreement lateral inhibitory mechanisms contribute to surround suppression, but little direct experimental evidence that intracortical inhibition plays a major role. Two intracellular recording studies of V1, one using naturalistic stimuli (Haider et al., 2010), the other sinusoidal gratings (Ozeki et al., 2009), sought to identify the causes of reduced activity in V1 with increasing stimulus size, a hallmark of surround suppression. The former attributed this effect to increased inhibition, the latter to largely balanced withdrawal of excitation and inhibition. In rodent primary somatosensory barrel cortex, multi-whisker responses are generally weaker than single whisker responses, suggesting multi-whisker stimulation engages similar surround suppressive mechanisms. The origins of suppression in S1 remain elusive: studies have implicated brainstem lateral/internuclear interactions and both thalamic and cortical inhibition. Although the anatomical organization and instantiation of surround suppression in the visual and somatosensory systems differ, we consider the idea that one common function of surround suppression, in both modalities, is to remove the statistical redundancies associated with natural stimuli by increasing the sparseness or selectivity of sensory responses.

Synaptic and network mechanisms of sparse and reliable visual cortical activity during nonclassical receptive field stimulation.

During natural vision, the entire visual field is stimulated by images rich in spatiotemporal structure. Although many visual system studies restrict stimuli to the classical receptive field (CRF), it is known that costimulation of the CRF and the surrounding nonclassical receptive field (nCRF) increases neuronal response sparseness. The cellular and network mechanisms underlying increased response sparseness remain largely unexplored. Here we show that combined CRF + nCRF stimulation increases the sparseness, reliability, and precision of spiking and membrane potential responses in classical regular spiking (RS(C)) pyramidal neurons of cat primary visual cortex. Conversely, fast-spiking interneurons exhibit increased activity and decreased selectivity during CRF + nCRF stimulation. The increased sparseness and reliability of RS(C) neuron spiking is associated with increased inhibitory barrages and narrower visually evoked synaptic potentials. Our experimental observations were replicated with a simple computational model, suggesting that network interactions among neuronal subtypes ultimately sharpen recurrent excitation, producing specific and reliable visual responses.