Tonotopic organization of the auditory cortex has been extensively studied in many mammalian species using various methodologies and physiological preparations. Tonotopy mapping in primates, however, is more limited due to constraints such as cortical folding, use of anesthetized subjects, and mapping methodology. Here we applied a combination of through-skull and through-window intrinsic optical signal imaging, wide-field calcium imaging, and neural probe recording techniques in awake marmosets (Callithrix jacchus), a New World monkey with most of its auditory cortex located on a flat brain surface. Coarse tonotopic gradients, including a recently described rostral-temporal (RT) to parabelt gradient, were revealed by the through-skull imaging of intrinsic optical signals and were subsequently validated by single-unit recording. Furthermore, these tonotopic gradients were observed with more detail through chronically implanted cranial windows with additional verifications on the experimental design. Moreover, the tonotopy mapped by the intrinsic-signal imaging methods was verified by wide-field calcium imaging in an AAV-GCaMP labeled subject. After these validations and with further effort to expand the field of view more rostrally in both windowed and through-skull subjects, an additional putative tonotopic gradient was observed more rostrally to the area RT, which has not been previously described by the standard model of tonotopic organization of the primate auditory cortex. Together, these results provide the most comprehensive data of tonotopy mapping in an awake primate species with unprecedented coverage and details in the rostral proportion and support a caudal-rostrally arranged mesoscale organization of at least three repeats of functional gradients in the primate auditory cortex, similar to the ventral stream of primate visual cortex.

Human speech and animal vocalizations are important for social communication and animal survival. Neurons in the auditory pathway are responsive to a range of sounds, from elementary sound features to complex acoustic sounds. For social communication, responses to distinct patterns of vocalization are usually highly specific to an individual conspecific call, in some species. This includes the specificity of sound patterns and embedded biological information. We conducted single-unit recordings in the amygdala of awake marmosets and presented calls used in marmoset communication, calls of other species and calls from specific marmoset individuals. We found that some neurons (47/262) in the amygdala distinguished \'Phee\' calls from vocalizations of other animals and other types of marmoset vocalizations. Interestingly, a subset of Phee-responsive neurons (22/47) also exhibited selectivity to one out of the three Phees from two different \'caller\' marmosets. Our findings suggest that, while it has traditionally been considered the key structure in the limbic system, the amygdala also represents a critical stage of socially relevant auditory perceptual processing.

Recent neuroimaging studies in humans have reported distinct temporal dynamics of gyri and sulci, which may be associated with putative functions of cortical gyrification. However, the complex folding patterns of the human cortex make it difficult to explain temporal patterns of gyrification. In this study, we used the common marmoset as a simplified model to examine the temporal characteristics and compare them with the complex gyrification of humans. Using a brain-inspired deep neural network, we obtained reliable temporal-frequency fingerprints of gyri and sulci from the awake rs-fMRI data of marmosets and humans. Notably, the temporal fingerprints of one region successfully classified the gyrus/sulcus of another region in both marmosets and humans. Additionally, the temporal-frequency fingerprints were remarkably similar in both species. We then analyzed the resulting fingerprints in several domains and adopted the Wavelet Transform Coherence approach to characterize the gyro-sulcal coupling patterns. In both humans and marmosets, sulci exhibited higher frequency bands than gyri, and the two were temporally coupled within the same range of phase angles. This study supports the notion that gyri and sulci possess unique and evolutionarily conserved features that are consistent across functional areas, and advances our understanding of the functional role of cortical gyrification.

As science and technology evolve, there is an increasing need for promotion of international scientific exchange. Collaborations, while offering substantial opportunities for scientists and benefit to society, also present challenges for those working with animal models, such as non-human primates (NHPs). Diversity in regulation of animal research is sometimes mistaken for the absence of common international welfare standards. Here, the ethical and regulatory protocols for 13 countries that have guidelines in place for biomedical research involving NHPs were assessed with a focus on neuroscience. Review of the variability and similarity in trans-national NHP welfare regulations extended to countries in Asia, Europe and North America. A tabulated resource was established to advance solution-oriented discussions and scientific collaborations across borders. Our aim is to better inform the public and other stakeholders. Through cooperative efforts to identify and analyze information with reference to evidence-based discussion, the proposed key ingredients may help to shape and support a more informed, open framework. This framework and resource can be expanded further for biomedical research in other countries.

The amygdala is an important hub for regulating emotions and is involved in the pathophysiology of many mental diseases, such as depression and anxiety. Meanwhile, the endocannabinoid system plays a crucial role in regulating emotions and mainly functions through the cannabinoid type-1 receptor (CB1R), which is strongly expressed in the amygdala of non-human primates (NHPs). However, it remains largely unknown how the CB1Rs in the amygdala of NHPs regulate mental diseases. Here, we investigated the role of CB1R by knocking down the cannabinoid receptor 1 (CNR1) gene encoding CB1R in the amygdala of adult marmosets through regional delivery of AAV-SaCas9-gRNA. We found that CB1R knockdown in the amygdala induced anxiety-like behaviors, including disrupted night sleep, agitated psychomotor activity in new environments, and reduced social desire. Moreover, marmosets with CB1R-knockdown had up-regulated plasma cortisol levels. These results indicate that the knockdown of CB1Rs in the amygdala induces anxiety-like behaviors in marmosets, and this may be the mechanism underlying the regulation of anxiety by CB1Rs in the amygdala of NHPs.

Hearing plays an important role in our ability to control voice, and perturbations in auditory feedback result in compensatory changes in vocal production. The auditory cortex (AC) has been proposed as an important mediator of this behavior, but causal evidence is lacking. We tested this in an animal model, hypothesizing that AC is necessary for vocal self-monitoring and feedback-dependent control, and that altering activity in AC during vocalization will interfere with vocal control.
We implanted two marmoset monkeys (Callithrix jacchus) with bilateral AC electrode arrays. Acoustic signals were recorded from vocalizing marmosets while altering vocal feedback or electrically stimulating AC during random subsets of vocalizations. Feedback was altered by real-time frequency shifts and presented through headphones and electrical stimulation delivered to individual electrodes. We analyzed recordings to measure changes in vocal acoustics during shifted feedback and stimulation, and to determine their interaction. Results were correlated with the location and frequency tuning of stimulation sites.
Consistent with previous results, we found electrical stimulation alone evoked changes in vocal production. Results were stronger in the right hemisphere, but decreased with lower currents or repeated stimulation. Simultaneous stimulation and shifted feedback significantly altered vocal control for a subset of sites, decreasing feedback compensation at some and increasing it at others. Inhibited compensation was more likely at sites closer to vocal frequencies.
Results provide causal evidence that the AC is involved in feedback-dependent vocal control, and that it is sufficient and may also be necessary to drive changes in vocal production.
N/A Laryngoscope, 133:1-10, 2023.

Our brains constantly generate predictions of sensory input that are compared with actual inputs, propagate the prediction-errors through a hierarchy of brain regions, and subsequently update the internal predictions of the world. However, the essential feature of predictive coding, the notion of hierarchical depth and its neural mechanisms, remains largely unexplored. Here, we investigated the hierarchical depth of predictive auditory processing by combining functional magnetic resonance imaging (fMRI) and high-density whole-brain electrocorticography (ECoG) in marmoset monkeys during an auditory local-global paradigm in which the temporal regularities of the stimuli were designed at two hierarchical levels. The prediction-errors and prediction updates were examined as neural responses to auditory mismatches and omissions. Using fMRI, we identified a hierarchical gradient along the auditory pathway: midbrain and sensory regions represented local, shorter-time-scale predictive processing followed by associative auditory regions, whereas anterior temporal and prefrontal areas represented global, longer-time-scale sequence processing. The complementary ECoG recordings confirmed the activations at cortical surface areas and further differentiated the signals of prediction-error and update, which were transmitted via putative bottom-up γ and top-down β oscillations, respectively. Furthermore, omission responses caused by absence of input, reflecting solely the two levels of prediction signals that are unique to the hierarchical predictive coding framework, demonstrated the hierarchical top-down process of predictions in the auditory, temporal, and prefrontal areas. Thus, our findings support the hierarchical predictive coding framework, and outline how neural networks and spatiotemporal dynamics are used to represent and arrange a hierarchical structure of auditory sequences in the marmoset brain.

The use of the common marmoset monkey (Callithrix jacchus) for neuroscientific research has grown markedly in the last decade. Magnetic resonance imaging (MRI) has played a significant role in establishing the extent of comparability of marmoset brain architecture with the human brain and brains of other preclinical species (eg, macaques and rodents). As a non-invasive technique, MRI allows for the flexible acquisition of the same sequences across different species in vivo, including imaging of whole-brain functional topologies not possible with more invasive techniques. Being one of the smallest New World primates, the marmoset may be an ideal nonhuman primate species to study with MRI. As primates, marmosets have an elaborated frontal cortex with features analogous to the human brain, while also having a small enough body size to fit into powerful small-bore MRI systems typically employed for rodent imaging; these systems offer superior signal strength and resolution. Further, marmosets have a rich behavioral repertoire uniquely paired with a lissencephalic cortex (like rodents). This smooth cortical surface lends itself well to MRI and also other invasive methodologies. With the advent of transgenic modification techniques, marmosets have gained significant traction as a powerful complement to canonical mammalian modelling species. Marmosets are poised to make major contributions to preclinical investigations of the pathophysiology of human brain disorders as well as more basic mechanistic explorations of the brain. The goal of this article is to provide an overview of the practical aspects of implementing MRI and fMRI in marmosets (both under anesthesia and fully awake) and discuss the development of resources recently made available for marmoset imaging.

The physiological characteristics of the marmoset second visual area (V2) are poorly understood compared with those of the primary visual area (V1). In this study, we observed the physiological response characteristics of V2 neurons in four healthy adult marmosets using intracortical tungsten microelectrodes. We recorded 110 neurons in area V2, with receptive fields located between 8° and 15° eccentricity. Most (88.2%) of these neurons were orientation selective, with half-bandwidths typically ranging between 10° and 30°. A significant proportion of neurons (28.2%) with direction selectivity had a direction index greater than 0.5. The vast majority of V2 neurons had separable spatial frequency and temporal frequency curves and, according to this criterion, they were not speed selective. The basic functional response characteristics of neurons in area V2 resemble those found in area V1. Our findings show that area V2 together with V1 are important in primate visual processing, especially in locating objects in space and in detecting an object\'s direction of motion. The methods used in this study were approved by the Monash University Animal Ethics Committee, Australia (MARP 2009-2011) in 2009.

Multi-modal neuroimaging projects such as the Human Connectome Project (HCP) and UK Biobank are advancing our understanding of human brain architecture, function, connectivity, and their variability across individuals using high-quality non-invasive data from many subjects. Such efforts depend upon the accuracy of non-invasive brain imaging measures. However, \'ground truth\' validation of connectivity using invasive tracers is not feasible in humans. Studies using nonhuman primates (NHPs) enable comparisons between invasive and non-invasive measures, including exploration of how \"functional connectivity\" from fMRI and \"tractographic connectivity\" from diffusion MRI compare with long-distance connections measured using tract tracing. Our NonHuman Primate Neuroimaging & Neuroanatomy Project (NHP_NNP) is an international effort (6 laboratories in 5 countries) to: (i) acquire and analyze high-quality multi-modal brain imaging data of macaque and marmoset monkeys using protocols and methods adapted from the HCP; (ii) acquire quantitative invasive tract-tracing data for cortical and subcortical projections to cortical areas; and (iii) map the distributions of different brain cell types with immunocytochemical stains to better define brain areal boundaries. We are acquiring high-resolution structural, functional, and diffusion MRI data together with behavioral measures from over 100 individual macaques and marmosets in order to generate non-invasive measures of brain architecture such as myelin and cortical thickness maps, as well as functional and diffusion tractography-based connectomes. We are using classical and next-generation anatomical tracers to generate quantitative connectivity maps based on brain-wide counting of labeled cortical and subcortical neurons, providing ground truth measures of connectivity. Advanced statistical modeling techniques address the consistency of both kinds of data across individuals, allowing comparison of tracer-based and non-invasive MRI-based connectivity measures. We aim to develop improved cortical and subcortical areal atlases by combining histological and imaging methods. Finally, we are collecting genetic and sociality-associated behavioral data in all animals in an effort to understand how genetic variation shapes the connectome and behavior.