Awake Macaque Monkeys Research Articles

Inserting a Neuropixels probe into awake monkey cortex: two probes, two methods

BackgroundNeuropixels probes have revolutionized neurophysiological studies in the rodent, but inserting these probes through the much thicker primate dura remains a challenge. New methodsHere we describe two methods we have developed for the insertion of two types of Neuropixels probes acutely into the awake macaque monkey cortex. For the fine rodent probe (Neuropixels 1.0, IMEC), which is unable to pierce native primate dura, we developed a dural-eyelet method to insert the probe repeatedly without breakage. For the thicker short NHP probe (Neuropixels NP1010), we developed an artificial dura system to insert the probe. Results and comparison with existing methodsWe have now conducted successful experiments in 3 animals across 7 recording chambers with the procedures described here and have achieved recordings with similar yields over several months in each case. ConclusionWe hope that our hardware, surgical preparation, methods for insertion and methods for removal of broken probe parts are of value to primate physiologists everywhere.

Intracranial EEG Recordings of High-Frequency Activity From a Wireless Implantable BMI Device in Awake Nonhuman Primates.

Wireless implantable brain machine interfaces (BMIs) are a promising tool to restore communication and motor functions for individuals with severe motor disability. Prior to clinical application, recording performance must be sufficiently confirmed by animal experiments. In this paper, we aimed to evaluate the performance of a novel BMI wireless device for recording brain activity in two nonhuman primates. We customized a wireless device for implantable BMIs for clinical application. We used a battery instead of a wireless power charging system. Thirty-two electrodes were subdurally implanted over the left temporoparietal cortex. We evaluated the recording performance of the wireless device by auditory steady-state responses (ASSRs) and ketamine-induced responses. The devices successfully recorded broadband oscillatory activities up to the high-frequency band from the temporal cortex in two awake macaque monkeys. Spectral analysis of raw signals demonstrated that the devices detected characteristic results of a 40-Hz ASSR and prominent high-frequency band activity induced by ketamine injection. We confirmed the functionality of the wireless device in recording and transmitting electrocorticography (ECoG) signals with both millisecond precision and recording stability. These results provide confidence that this wireless device can be a translational tool for other fundamental neuroscientific studies in free-moving models.

Suppressive effects of ketamine on auditory steady-state responses in intact, awake macaques: A non-human primate model of schizophrenia

Auditory steady-state responses (ASSRs) are recurrent neural activities entrained to regular cyclic auditory stimulation. ASSRs are altered in individuals with schizophrenia, and may be related to hypofunction of the N-methyl-D-aspartate (NMDA) glutamate receptor. Noncompetitive NMDA receptor antagonists, including ketamine, have been used in ASSR studies of rodent models of schizophrenia. Although animal studies using non-human primates are required to complement rodent studies, the effects of ketamine on ASSRs are unknown in intact awake non-human primates. In this study, after administration of vehicle or ketamine, click trains at 20–83.3 Hz were presented to elicit ASSRs during recording of electroencephalograms in intact, awake macaque monkeys. The results indicated that ASSRs quantified by event-related spectral perturbation and inter-trial coherence were maximal at 83.3 Hz after vehicle administration, and that ketamine reduced ASSRs at 58.8 and 83.3 Hz, but not at 20 and 40 Hz. The present results demonstrated a reduction of ASSRs by the NMDA receptor antagonist at optimal frequencies with maximal responses in intact, awake macaques, comparable to ASSR reduction in patients with schizophrenia. These findings suggest that ASSR can be used as a neurophysiological biomarker of the disturbance of gamma-oscillatory neural circuits in this ketamine model of schizophrenia using intact, awake macaques. Thus, this model with ASSRs would be useful in the investigation of human brain pathophysiology as well as in preclinical translational research.

Decoding Pixel-Level Image Features From Two-Photon Calcium Signals of Macaque Visual Cortex.

Images of visual scenes comprise essential features important for visual cognition of the brain. The complexity of visual features lies at different levels, from simple artificial patterns to natural images with different scenes. It has been a focus of using stimulus images to predict neural responses. However, it remains unclear how to extract features from neuronal responses. Here we address this question by leveraging two-photon calcium neural data recorded from the visual cortex of awake macaque monkeys. With stimuli including various categories of artificial patterns and diverse scenes of natural images, we employed a deep neural network decoder inspired by image segmentation technique. Consistent with the notation of sparse coding for natural images, a few neurons with stronger responses dominated the decoding performance, whereas decoding of ar tificial patterns needs a large number of neurons. When natural images using the model pretrained on artificial patterns are decoded, salient features of natural scenes can be extracted, as well as the conventional category information. Altogether, our results give a new perspective on studying neural encoding principles using reverse-engineering decoding strategies.

Interaction between steady-state visually evoked potentials at nearby flicker frequencies

Steady-state visually evoked potential (SSVEP) studies routinely employ simultaneous presentation of two temporally modulated stimuli, with SSVEP amplitude modulations serving to index top-down cognitive processes. However, the nature of SSVEP amplitude modulations as a function of competing temporal frequency (TF) has not been systematically studied, especially in relation to the normalization framework which has been extensively used to explain visual responses to multiple stimuli. We recorded spikes and local field potential (LFP) from the primary visual cortex (V1) as well as EEG from two awake macaque monkeys while they passively fixated plaid stimuli with components counterphasing at different TFs. We observed asymmetric SSVEP response suppression by competing TFs (greater suppression for lower TFs), which further depended on the relative orientations of plaid components. A tuned normalization model, adapted to SSVEP responses, provided a good account of the suppression. Our results provide new insights into processing of temporally modulated visual stimuli.

Two-photon imaging of V1 responses to complex stimulus patterns in awake macaque monkeys

Two-photon imaging of V1 responses to complex stimulus patterns in awake macaque monkeys

Second type of criticality in the brain uncovers rich multiple-neuron dynamics

Cortical networks that have been found to operate close to a critical point exhibit joint activations of large numbers of neurons. However, in motor cortex of the awake macaque monkey, we observe very different dynamics: massively parallel recordings of 155 single-neuron spiking activities show weak fluctuations on the population level. This a priori suggests that motor cortex operates in a noncritical regime, which in models, has been found to be suboptimal for computational performance. However, here, we show the opposite: The large dispersion of correlations across neurons is the signature of a second critical regime. This regime exhibits a rich dynamical repertoire hidden from macroscopic brain signals but essential for high performance in such concepts as reservoir computing. An analytical link between the eigenvalue spectrum of the dynamics, the heterogeneity of connectivity, and the dispersion of correlations allows us to assess the closeness to the critical point.

Microdialysis in awake macaque monkeys for central nervous system pharmacokinetics.

BackgroundThe brain bioavailability of novel small molecules developed to address central nervous system disease is classically documented through ex vivo or in vivo analyses conducted in rodent models. Data acquired in rodent models are, however, not easily transferrable to human as the pharmacokinetic and pharmacodynamics profiles of the species are quite different.MethodsUsing drugs selected for their differential transport across the blood‐brain barrier, we here demonstrate the feasibility of brain microdialysis in normal vigil macaque monkey by measuring brain extracellular fluid bioavailability of carbamazepine, digoxin, oxycodone, and quinidine.ResultsAll drugs, but digoxin, were found in dialysate samples. Drugs that are substrate of P‐glycoprotein show a difference of bioavailability or brain pharmacokinetic parameters between rodents and primates.ConclusionData suggest that brain microdialysis in vigil macaque monkey, the species of choice for classic pharmacokinetic/pharmacodynamics studies could help predicting human brain bioavailability of a small molecule depending on the protein involved in the efflux transport from the brain.

Monkey-MIMMS: Towards Automated Cellular Resolution Large- Scale Two-Photon Microscopy In The Awake Macaque Monkey.

The size and curvature of the macaque brain present challenges for two photon laser scanning microscopy (2P-LSM). General access to the cortex requires 5-axis positioning over a range of motion wider than existing designs offer. In addition, movement artifacts due to physiological pulsations and bodily movement present particular challenges. We present a microscope and implant platform that allows for repeatable, motorized positioning and stable imaging at any point on the dorsal convexity of macaque cortex. While testing the system to image neurons expressing fluorescent proteins in an awake macaque, motion artifacts were limited to several microns.

Convolutional neural network models of V1 responses to complex patterns.

In this study, we evaluated the convolutional neural network (CNN) method for modeling V1 neurons of awake macaque monkeys in response to a large set of complex pattern stimuli. CNN models outperformed all the other baseline models, such as Gabor-based standard models for V1 cells and various variants of generalized linear models. We then systematically dissected different components of the CNN and found two key factors that made CNNs outperform other models: thresholding nonlinearity and convolution. In addition, we fitted our data using a pre-trained deep CNN via transfer learning. The deep CNN's higher layers, which encode more complex patterns, outperformed lower ones, and this result was consistent with our earlier work on the complexity of V1 neural code. Our study systematically evaluates the relative merits of different CNN components in the context of V1 neuron modeling.

Neuronal Correlates of Auditory Streaming in Monkey Auditory Cortex for Tone Sequences without Spectral Differences

This study finds a neuronal correlate of auditory perceptual streaming in the primary auditory cortex for sequences of tone complexes that have the same amplitude spectrum but a different phase spectrum. Our finding is based on microelectrode recordings of multiunit activity from 270 cortical sites in three awake macaque monkeys. The monkeys were presented with repeated sequences of a tone triplet that consisted of an A tone, a B tone, another A tone and then a pause. The A and B tones were composed of unresolved harmonics formed by adding the harmonics in cosine phase, in alternating phase, or in random phase. A previous psychophysical study on humans revealed that when the A and B tones are similar, humans integrate them into a single auditory stream; when the A and B tones are dissimilar, humans segregate them into separate auditory streams. We found that the similarity of neuronal rate responses to the triplets was highest when all A and B tones had cosine phase. Similarity was intermediate when the A tones had cosine phase and the B tones had alternating phase. Similarity was lowest when the A tones had cosine phase and the B tones had random phase. The present study corroborates and extends previous reports, showing similar correspondences between neuronal activity in the primary auditory cortex and auditory streaming of sound sequences. It also is consistent with Fishman’s population separation model of auditory streaming.

Long-Term Two-Photon Imaging in Awake Macaque Monkey

Successful application of two-photon imaging withgenetic tools in awake macaque monkeys will enable fundamental advances in our understanding of higher cognitive function at the level of molecular and neuronal circuits. Here we report techniques for long-term two-photon imaging in awake macaque monkeys. Using genetically encoded indicators including GCaMP5 and GCaMP6s delivered by AAV2/1 into the visual cortex, we demonstrate that high-quality two-photon imaging of large neuronal populations can be achieved and maintained in awake monkeys for months. Simultaneous intracellular recording and two-photon calcium imaging confirm that fluorescence activity is linearly proportional to neuronal spiking activity across a wide range of firing rates (10Hz to 150Hz). By providing two-photon imaging access to cortical neuronal populations at single-cell or single dendritic spine resolution in awake monkeys, the techniques reported can help bridge the use of modern genetic and molecular tools and the study of higher cognitive function.

Spectral and spatial tuning of onset and offset response functions in auditory cortical fields A1 and CL of rhesus macaques.

The mammalian auditory cortex is necessary for spectral and spatial processing of acoustic stimuli. Most physiological studies of single neurons in the auditory cortex have focused on the onset and sustained portions of evoked responses, but there have been far fewer studies on the relationship between onset and offset responses. In the current study, we compared spectral and spatial tuning of onset and offset responses of neurons in primary auditory cortex (A1) and the caudolateral (CL) belt area of awake macaque monkeys. Several different metrics were used to determine the relationship between onset and offset response profiles in both frequency and space domains. In the frequency domain, a substantial proportion of neurons in A1 and CL displayed highly dissimilar best stimuli for onset- and offset-evoked responses, although even for these neurons, there was usually a large overlap in the range of frequencies that elicited onset, and offset responses and distributions of tuning overlap metrics were mostly unimodal. In the spatial domain, the vast majority of neurons displayed very similar best locations for onset- and offset-evoked responses, along with unimodal distributions of all tuning overlap metrics considered. Finally, for both spectral and spatial tuning, a slightly larger fraction of neurons in A1 displayed nonoverlapping onset and offset response profiles, relative to CL, which supports hierarchical differences in the processing of sounds in the two areas. However, these differences are small compared with differences in proportions of simple cells (low overlap) and complex cells (high overlap) in primary and secondary visual areas.NEW & NOTEWORTHY In the current study, we examine the relationship between the tuning of neural responses evoked by the onset and offset of acoustic stimuli in the primary auditory cortex, as well as a higher-order auditory area-the caudolateral belt field-in awake rhesus macaques. In these areas, the relationship between onset and offset response profiles in frequency and space domains formed a continuum, ranging from highly overlapping to highly nonoverlapping.

A systematic analysis of neurons with large somatosensory receptive fields covering multiple body regions in the secondary somatosensory area of macaque monkeys.

Previous neurophysiological studies performed in macaque monkeys have revealed complex somatosensory responses in the secondary somatosensory area (SII), such as large receptive fields (RFs), as well as bilateral ones. However, systematic analyses of neurons with large RFs have not been performed. In the present study, we recorded single-unit activities in SII of awake macaque monkeys to investigate systematically large RFs by dividing the whole body into four body regions (head, trunk, forelimb, and hindlimb). Recorded neurons were classified into two types, according to whether the RFs were confined to one body region: single (n = 817) and combined (n = 282) body-region types. These two types were distinct in terms of the percentage of bilateral RFs: 55% in the single-region type and 90% in the combined type, demonstrating that two types of RF enlargement occur simultaneously in the combined type, namely, RF convergence from different body regions and RF convergence from both hemibodies. Among the combined-type RFs, two tendencies of RF convergence were found: 1) the distal parts of the limbs (i.e., hand and foot) and the mouth are interconnected, and 2) the trunk RFs extend continuously toward the distal parts of the limb and head to cover the entire body surface. Our distribution analysis on unfolded maps clarified that neurons having RFs with these two tendencies were distributed within specific subregions in SII.

Spatial structure of neuronal receptive field in awake monkey secondary visual cortex (V2)

Visual processing depends critically on the receptive field (RF) properties of visual neurons. However, comprehensive characterization of RFs beyond the primary visual cortex (V1) remains a challenge. Here we report fine RF structures in secondary visual cortex (V2) of awake macaque monkeys, identified through a projection pursuit regression analysis of neuronal responses to natural images. We found that V2 RFs could be broadly classified as V1-like (typical Gabor-shaped subunits), ultralong (subunits with high aspect ratios), or complex-shaped (subunits with multiple oriented components). Furthermore, single-unit recordings from functional domains identified by intrinsic optical imaging showed that neurons with ultralong RFs were primarily localized within pale stripes, whereas neurons with complex-shaped RFs were more concentrated in thin stripes. Thus, by combining single-unit recording with optical imaging and a computational approach, we identified RF subunits underlying spatial feature selectivity of V2 neurons and demonstrated the functional organization of these RF properties.

Facilitative effect of repetitive presentation of one stimulus on cortical responses to other stimuli in macaque monkeys--a possible neural mechanism for mismatch negativity.

The event‐related potential ‘mismatch negativity’ (MMN) is an indicator of a perceiver's ability to detect deviations in sensory signal streams. MMN and its homologue in animals, mismatch activity (MMA), are differential neural responses to a repeatedly presented stimulus and a subsequent deviant stimulus (oddball). Because neural mechanisms underlying MMN and MMA remain unclear, there is a controversy as to whether MMN and MMA arise solely from stimulus‐specific adaptation (SSA), in which the response to a stimulus cumulatively attenuates with its repetitive presentation. To address this issue, we used electrocorticography and the auditory roving‐oddball paradigm in two awake macaque monkeys. We examined the effect of stimulus repetition number on MMA and on responses to repeated stimuli and oddballs across the cerebral cortex in the time–frequency domain. As the repetition number increased, MMA spread across the temporal, frontal and parietal cortices, and each electrode yielded a larger MMA. Surprisingly, this increment in MMA largely depended on response augmentation to the oddball rather than on SSA to the repeated stimulus. Following sufficient repetition, the oddball evoked a spectral power increment in some electrodes on the frontal cortex that had shown no power increase to the stimuli with less or no preceding repetition. We thereby revealed that repetitive presentation of one stimulus not only leads to SSA but also facilitates the cortical response to oddballs involving a wide range of cortical regions. This facilitative effect might underlie the generation of MMN‐like scalp potentials in macaques that potentially shares similar neural mechanisms with MMN in humans.

Vision for Prehension in the Medial Parietal Cortex.

In the last 2 decades, the medial posterior parietal area V6A has been extensively studied in awake macaque monkeys for visual and somatosensory properties and for its involvement in encoding of spatial parameters for reaching, including arm movement direction and amplitude. This area also contains populations of neurons sensitive to grasping movements, such as wrist orientation and grip formation. Recent work has shown that V6A neurons also encode the shape of graspable objects and their affordance. In other words, V6A seems to encode object visual properties specifically for the purpose of action, in a dynamic sequence of visuomotor transformations that evolve in the course of reach-to-grasp action.We propose a model of cortical circuitry controlling reach-to-grasp actions, in which V6A acts as a comparator that monitors differences between current and desired hand positions and configurations. This error signal could be used to continuously update the motor output, and to correct reach direction, hand orientation, and/or grip aperture as required during the act of prehension.In contrast to the generally accepted view that the dorsomedial component of the dorsal visual stream encodes reaching, but not grasping, the functional properties of V6A neurons strongly suggest the view that this area is involved in encoding all phases of prehension, including grasping.

Neuronal response properties in area MT of an awake amblyopic macaque monkey

Neuronal response properties in area MT of an awake amblyopic macaque monkey

Is the mirror cracked?

Few recent discoveries in the field of neuroscience have given rise to quite as much interest and controversy as that of mirror neurons, first reported by Giacomo Rizzolatti and his colleagues at the University of Parma in 1992. The first full description of these intriguing entities was published 4 years later in Brain (Gallese et al. , 1996). These authors described the activity of a group of neurons recorded in the ventral premotor cortex of the awake macaque monkey, in a subdivision referred to as F(rontal)5. These F5 neurons fired during particular types of grasp carried out by the monkey; there was nothing surprising about that, since this area had already been identified as a cortical area that was grasp-related. The striking finding was that the same neurons also discharged in a highly reproducible way as the monkey observed a human experimenter making a similar grasp. There was no evidence that this discharge was related to covert movements made by the monkey as it watched: the discharge was clearly evoked by the action of another individual. This fascinating discovery has now been confirmed by several other laboratories (Kilner and Lemon, 2013). Gregory Hickok’s book on this controversial subject carries an intentionally mischievous title: in fact, there is nothing mythological about the existence of mirror neurons (Arbib, 2015), and their discovery has completely changed the way we think about how our own brain interacts with that of others. However, there are plenty of stories about how mirror neurons might function, and in particular how experimental details of mirror neurons acquired in monkeys might illuminate the function of a ‘mirror neuron system’ in humans. Hickok’s well-known difficulties with the proposed functions of a human mirror neuron system stem from two major sets of issues. One is the tension introduced by linking …

Visual Responsiveness of Neurons in the Secondary Somatosensory Area and its Surrounding Parietal Operculum Regions in Awake Macaque Monkeys.

Previous neurophysiological studies performed in macaque monkeys have shown that the secondary somatosensory cortex (SII) is essentially engaged in the processing of somatosensory information and no other sensory input has been reported. In contrast, recent human brain-imaging studies have revealed the effects of visual and auditory stimuli on SII activity, which suggest multisensory integration in the human SII. To determine whether multisensory responses of the SII also exist in nonhuman primates, we recorded single-unit activity in response to visual and auditory stimuli from the SII and surrounding regions in 8 hemispheres from 6 awake monkeys. Among 1157 recorded neurons, 306 neurons responded to visual stimuli. These visual neurons usually responded to rather complex stimuli, such as stimulation of the peripersonal space (40.5%), observation of human action (29.1%), and moving-object stimulation outside the monkey's reach (23.9%). We occasionally applied auditory stimuli to visual neurons and found 10 auditory-responsive neurons that exhibited somatosensory responses. The visual neurons were distributed continuously along the lateral sulcus covering the entire SII, along with other somatosensory neurons. These results highlight the need to investigate novel functional roles—other than somesthetic sensory processing—of the SII.