electroencephalography-measurements-awake-marmosets-listening-to

Electroencephalography Measurements in Awake Marmosets Listening to Conspecific Vocalizations

Vocal communication plays a crucial role in the social interactions of primates, particularly in survival and social organization. Humans have developed a ...

Kyoto University

Common marmosets have been used extensively in biomedical research and the recent advent of techniques to generate transgenic marmosets has accelerated the use of this model. New methods that efficiently assess the degree of cognitive function in common marmosets are needed in order to establish their suitability as non-human primate models of higher brain function disorders. Here, we have developed a new apparatus suitable for testing the cognitive functions of common marmosets. Utilizing a mini laptop PC with a touch-sensitive screen as the main component, the apparatus is small and lightweight and can be easily attached to the home cages. The ease of designing and testing new paradigms with the flexible software is another advantage of this system. We have tested visual discrimination and its reversal tasks using this apparatus and confirmed its efficacy.

Development of a compact and general-purpose experimental apparatus with a touch-sensitive screen for use in evaluating cognitive functions in common marmosets.

We investigated the characteristics of serial order learning in common marmosets (Callithrix jacchus). Five marmosets were trained in a sequential responding task in which they were required to touch four graphic patterns in a given order (A→B→C→D) to obtain a reward. All five marmosets learned the task with over 65% accuracy. Shuffling the positions of B, C, and D immediately after the marmoset had correctly identified and selected the first stimulus (A) either decreased accuracy or lengthened response latency for the second stimulus (B). These results suggest that the marmosets planned the response to the second stimulus before they touched the first stimulus. In addition, when we presented a pair of stimuli (AB, AC, AD, BC, BD, and CD pairs), the marmosets responded to the stimuli in the pair in the appropriate order, according to the learned order (A→B→C→D). The analyses of first and second response latencies clearly demonstrated both the first-item and missing-item effects in task performance. Our data provide direct evidence that marmosets can learn the relative order of the four stimuli in a sequential responding task.

Characteristics of serial order learning in common marmosets (Callithrix jacchus).

Detailed information about the characteristics of learning behavior in marmosets is useful for future marmoset research. We trained 42 marmosets in visual discrimination and reversal learning. All marmosets could learn visual discrimination, and all but one could complete reversal learning, though some marmosets failed to touch the visual stimuli and were screened out. In 87% of measurements, the final percentage of correct responses was over 95%. We quantified performance with two measures: onset trial and dynamic interval. Onset trial represents the number of trials that elapsed before the marmoset started to learn. Dynamic interval represents the number of trials from the start before reaching the final percentage of correct responses. Both measures decreased drastically as a result of the formation of discrimination learning sets. In reversal learning, both measures worsened, but the effect on onset trial was far greater. The effects of age and sex were not significant as far as we used adolescent or young adult marmosets. Unexpectedly, experimental circumstance (in the colony or isolator) had only a subtle effect on performance. However, we found that marmosets from different families exhibited different learning process characteristics, suggesting some family effect on learning.

Individual variability in visual discrimination and reversal learning performance in common marmosets.

It remains unknown how single-shot anesthesia influences physical parameters, especially respiratory function and blood oxygen level of common marmosets (Callithrix jacchus) which came to be used for laboratory research.

Hypoxemia after single-shot anesthesia in common marmosets.

Learning and memory processes are similarly organized in humans and monkeys; therefore, monkeys can be ideal models for analyzing human aging processes and neurodegenerative diseases such as Alzheimer's disease. With the development of novel gene modification methods, common marmosets () have been suggested as an animal model for neurodegenerative diseases. Furthermore, the common marmoset's lifespan is relatively short, which makes it a practical animal model for aging. Working memory deficits are a prominent symptom of both dementia and aging, but no data are currently available for visual working memory in common marmosets. The delayed matching-to-sample task is a powerful tool for evaluating visual working memory in humans and monkeys; therefore, we developed a novel procedure for training common marmosets in such a task. Using visual discrimination and reversal tasks to direct the marmosets' attention to the physical properties of visual stimuli, we successfully trained 11 out of 13 marmosets in the initial stage of the delayed matching-to-sample task and provided the first available data on visual working memory in common marmosets. We found that the marmosets required many trials to initially learn the task (median: 1316 trials), but once the task was learned, the animals needed fewer trials to learn the task with novel stimuli (476 trials or fewer, with the exception of one marmoset). The marmosets could retain visual information for up to 16 s. Our novel training procedure could enable us to use the common marmoset as a useful non-human primate model for studying visual working memory deficits in neurodegenerative diseases and aging.

A Method to Train Marmosets in Visual Working Memory Task and Their Performance.

Pseudotyped lentiviral vectors give access to pathway-selective gene manipulation via retrograde transfer. Two types of such lentiviral vectors have been developed. One is the so-called NeuRet vector pseudotyped with fusion glycoprotein type E, which preferentially transduces neurons. The other is the so-called HiRet vector pseudotyped with fusion glycoprotein type B2, which permits gene transfer into both neurons and glial cells at the injection site. Although these vectors have been applied in many studies investigating neural network functions, it remains unclear which vector is more appropriate for retrograde gene delivery in the brain. To compare the gene transfer efficiency and inflammatory response of the NeuRet vs. HiRet vectors, each vector was injected into the striatum in macaque monkeys, common marmosets, and rats. It was revealed that retrograde gene delivery of the NeuRet vector was equal to or greater than that of the HiRet vector. Furthermore, inflammation characterized by microglial and lymphocytic infiltration occurred when the HiRet vector, but not the NeuRet vector, was injected into the primate brain. The present results indicate that the NeuRet vector is more suitable than the HiRet vector for retrograde gene transfer in the primate and rodent brains.

A note on retrograde gene transfer efficiency and inflammatory response of lentiviral vectors pseudotyped with FuG-E vs. FuG-B2 glycoproteins.

The selective serotonin reuptake inhibitor fluoxetine (FLX) is widely used to treat depression and anxiety disorders. Chronic FLX treatment reportedly induces cellular responses in the brain, including increased adult hippocampal and cortical neurogenesis and reversal of neuron maturation in the hippocampus, amygdala, and cortex. However, because most previous studies have used rodent models, it remains unclear whether these FLX-induced changes occur in the primate brain. To evaluate the effects of FLX in the primate brain, we used immunohistological methods to assess neurogenesis and the expression of neuronal maturity markers following chronic FLX treatment (3 mg/kg/day for 4 weeks) in adult marmosets (n = 3 per group). We found increased expression of doublecortin and calretinin, markers of immature neurons, in the hippocampal dentate gyrus of FLX-treated marmosets. Further, FLX treatment reduced parvalbumin expression and the number of neurons with perineuronal nets, which indicate mature fast-spiking interneurons, in the hippocampus, but not in the amygdala or cerebral cortex. We also found that FLX treatment increased the generation of cortical interneurons; however, significant up-regulation of adult hippocampal neurogenesis was not observed in FLX-treated marmosets. These results suggest that dematuration of hippocampal neurons and increased cortical neurogenesis may play roles in FLX-induced effects and/or side effects. Our results are consistent with those of previous studies showing hippocampal dematuration and increased cortical neurogenesis in FLX-treated rodents. In contrast, FLX did not affect hippocampal neurogenesis or dematuration of interneurons in the amygdala and cerebral cortex.

Fluoxetine-induced dematuration of hippocampal neurons and adult cortical neurogenesis in the common marmoset.

It is agreed upon that adult hippocampal neurogenesis (AHN) occurs in the dentate gyrus (DG) in rodents. However, the existence of AHN in humans, particularly in elderly individuals, remains to be determined. Recently, several studies reported that neural progenitor cells, neuroblasts, and immature neurons were detected in the hippocampus of elderly humans, based on the expressions of putative markers for these cells, claiming that this provides evidence of the persistence of AHN in humans. Herein, we briefly overview the phenomenon that we call "dematuration," in which mature neurons dedifferentiate to a pseudo-immature status and re-express the molecular markers of neural progenitor cells and immature neurons. Various conditions can easily induce dematuration, such as inflammation and hyper-excitation of neurons, and therefore, the markers for neural progenitor cells and immature neurons may not necessarily serve as markers for AHN. Thus, the aforementioned studies have not presented definitive evidence for the persistence of hippocampal neurogenesis throughout adult life in humans, and we would like to emphasize that those markers should be used cautiously when presented as evidence for AHN. Increasing AHN has been considered as a therapeutic target for Alzheimer's disease (AD); however, given that immature neuronal markers can be re-expressed in mature adult neurons, independent of AHN, in various disease conditions including AD, strategies to increase the expression of these markers in the DG may be ineffective or may worsen the symptoms of such diseases.

Expression of progenitor cell/immature neuron markers does not present definitive evidence for adult neurogenesis.

Anaesthesia is often required in common marmosets undergoing various procedures. The aim of this study was to evaluate anaesthetic and cardiopulmonary effects of alfaxalone, alfaxalone-ketamine and alfaxalone-butorphanol-medetomidine in common marmosets.

Evaluation of anaesthetic and cardiorespiratory effects after intramuscular administration of alfaxalone alone, alfaxalone-ketamine and alfaxalone-butorphanol-medetomidine in common marmosets (Callithrix jacchus).

For achieving retrograde gene transfer, we have so far developed two types of lentiviral vectors pseudotyped with fusion envelope glycoprotein, termed HiRet vector and NeuRet vector, consisting of distinct combinations of rabies virus and vesicular stomatitis virus glycoproteins. In the present study, we compared the patterns of retrograde transgene expression for the HiRet vs. NeuRet vectors by testing the cortical input system. These vectors were injected into the motor cortex in rats, marmosets, and macaques, and the distributions of retrograde labels were investigated in the cortex and thalamus. Our histological analysis revealed that the NeuRet vector generally exhibits a higher efficiency of retrograde gene transfer than the HiRet vector, though its capacity of retrograde transgene expression in the macaque brain is unexpectedly low, especially in terms of the intracortical connections, as compared to the rat and marmoset brains. It was also demonstrated that the NeuRet but not the HiRet vector displays sufficiently high neuron specificity and causes no marked inflammatory/immune responses at the vector injection sites in the primate (marmoset and macaque) brains. The present results indicate that the retrograde transgene efficiency of the NeuRet vector varies depending not only on the species but also on the input projections.

Retrograde Transgene Expression via Neuron-Specific Lentiviral Vector Depends on Both Species and Input Projections.

Auditory steady-state responses (ASSRs) are basic neural responses used to probe the ability of auditory circuits to produce synchronous activity to repetitive external stimulation. Reduced ASSR has been observed in patients with schizophrenia, especially at 40 Hz. Although ASSR is a translatable biomarker with a potential both in animal models and patients with schizophrenia, little is known about the features of ASSR in monkeys. Herein, we recorded the ASSR from humans, rhesus monkeys, and marmosets using the same method to directly compare the characteristics of ASSRs among the species. We used auditory trains on a wide range of frequencies to investigate the suitable frequency for ASSRs induction, because monkeys usually use stimulus frequency ranges different from humans for vocalization. We found that monkeys and marmosets also show auditory event-related potentials and phase-locking activity in gamma-frequency trains, although the optimal frequency with the best synchronization differed among these species. These results suggest that the ASSR could be a useful translational, cross-species biomarker to examine the generation of gamma-band synchronization in nonhuman primate models of schizophrenia.

Comparison of non-invasive, scalp-recorded auditory steady-state responses in humans, rhesus monkeys, and common marmosets.

Common marmosets usually give birth to twins and form a social group consisting of a breeding couple and pairs of same-aged siblings. The twins may engage in the first agonistic fights between them, twin-fights (TFs), during adolescence. This study investigated the TFs based on records accumulated in our captive colony over 12 years to elucidate the proximate causations that trigger the TFs. We aimed to determine whether the TF onset mainly depended on internal events (such as the onset of puberty) as previously suggested or external events (such as the birth of the younger siblings and the behavioral change of the group members). Although both events usually occur simultaneously, the birth control method (i.e., manipulation of ovulation and interbirth-intervals by prostaglandin administration to females) could temporally separate these events. A comparison of the onset day and occurrence rate with or without the birth control procedure revealed that TFs were triggered by a combination of internal and external events, that is, external events were the predominant triggers of TF, under the influence of internal events. The timing of TF onset was significantly delayed when the birth of the younger siblings was delayed and the twins grew older under the birth-controlled condition, suggesting that the birth of younger siblings and related behavioral changes of group members, as well as twins' developmental maturation, could trigger TF. Higher TF rates between same-sex twins were consistent with previous studies, reflecting the characteristics of same-sex directed aggression in callitrichines.

Article	Year
Development of a compact and general-purpose experimental apparatus with a touch-sensitive screen for use in evaluating cognitive functions in common marmosets. Journal of neuroscience methods\| PubMed ID: 21549751	2011
Characteristics of serial order learning in common marmosets (Callithrix jacchus). Journal of comparative psychology (Washington, D.C. : 1983)\| PubMed ID: 22468938	2012
Individual variability in visual discrimination and reversal learning performance in common marmosets. Neuroscience research\| PubMed ID: 25315397	2015
Hypoxemia after single-shot anesthesia in common marmosets. Journal of medical primatology\| PubMed ID: 28326553	2017
A Method to Train Marmosets in Visual Working Memory Task and Their Performance. Frontiers in behavioral neuroscience\| PubMed ID: 29615876	2018
A note on retrograde gene transfer efficiency and inflammatory response of lentiviral vectors pseudotyped with FuG-E vs. FuG-B2 glycoproteins. Scientific reports\| PubMed ID: 30837514	2019
Fluoxetine-induced dematuration of hippocampal neurons and adult cortical neurogenesis in the common marmoset. Molecular brain\| PubMed ID: 31383032	2019
Expression of progenitor cell/immature neuron markers does not present definitive evidence for adult neurogenesis. Molecular brain\| PubMed ID: 31823803	2019
Evaluation of anaesthetic and cardiorespiratory effects after intramuscular administration of alfaxalone alone, alfaxalone-ketamine and alfaxalone-butorphanol-medetomidine in common marmoset... Journal of medical primatology\| PubMed ID: 32654222	2020
Retrograde Transgene Expression via Neuron-Specific Lentiviral Vector Depends on Both Species and Input Projections. Viruses\| PubMed ID: 34372593	2021
Comparison of non-invasive, scalp-recorded auditory steady-state responses in humans, rhesus monkeys, and common marmosets. Scientific reports\| PubMed ID: 35654875	2022
Trigger of twin-fights in captive common marmosets. American journal of primatology\| PubMed ID: 37301733	2023

Miki Miwa

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