Pentylenetetrazole kindling induces dynamic changes in GAD65 expression in hippocampal somatostatin interneurons.

INTRODUCTION: One of the mechanisms of epileptgenesis is impairment of inhibitory neural circuits. Several studies have compared neural changes among subtypes of gamma-aminobutyric acid-related (GABAergic) neurons after acquired epileptic seizure. However, it is unclear that GABAergic neural modifications that occur during acquisition process of epileptic seizure. METHODS: Male rats were injected with pentylenetetrazole (PTZ kindling: n = 30) or saline (control: n = 15) every other day to observe the development of epileptic seizure stages. Two time points were identified: the point at which seizures were most difficult to induce, and the point at which seizures were most easy to induce. The expression of GABAergic neuron-related proteins in the hippocampus was immunohistochemically compared among GABAergic subtypes at each of these time points. RESULTS: Bimodal changes in seizure stages were observed in response to PTZ kindling. The increase of seizure stage was transiently suppressed after 8 or 10 injections, and then progressed again by the 16th injection. Based on these results, we defined 10 injections as a short-term injection period during which seizures are less likely to occur, and 20 injections as a long-term injection period during which continuous seizures are likely to occur. The immunohistochemical analysis showed that hippocampal glutamic acid decarboxylase 65 (GAD65) expression was increased after short-term kindling but unchanged after long-term kindling. Increased GAD65 expression was limited to somatostatin-positive (SOM+) cells among several GABAergic subtypes. By contrast, GAD, GABA, GABAAR α1, GABABR1, and VGAT cells showed no change following short- or long-term PTZ kindling. CONCLUSION: PTZ kindling induces bimodal changes in the epileptic seizure stage. Seizure stage is transiently suppressed after short-term PTZ injection with GAD65 upregulation in SOM+ cells. The seizure stage is progressed again after long-term PTZ injection with GAD65 reduction to baseline level.

Recruitment of the premotor cortex during arithmetic operations by the monkey.

Arithmetic operations are complex mental processes rooted in the abstract concept of numerosity. Despite the significance, the neural architecture responsible for these operations has remained largely uncharted. In this study, we explored the presence of specific neuronal activity in the dorsal premotor cortex of the monkey dedicated to numerical addition and subtraction. Our findings reveal that many of these neural activities undergo a transformation, shifting their coding from arithmetic to motor representations. These motor representations include information about which hand to use and the number of steps involved in the action. We consistently observed that cells related to the right-hand encoded addition, while those linked to the left-hand encoded subtraction, suggesting that arithmetic operations and motor commands are intertwining with each other. Furthermore, we used a multivariate decoding technique to predict the monkey's behaviour based on the activity of these arithmetic-related cells. The classifier trained to discern arithmetic operations, including addition and subtraction, not only predicted the arithmetic decisions but also the subsequent motor actions of the right and left-hand. These findings imply a cognitive extension of the motor cortex's function, where inherent neural systems are repurposed to facilitate arithmetic operations.

[Veronal, in Lord Edgware Dies: A Double-edged Sword of Sedation and Addiction].

Veronal, introduced during the early 20th century was widely used as a sleeping pill at the time, and this drug is mentioned in Agatha Christie's novels. Research has proved that in addition to their sedative effects, Veronal and other sleeping pills with similar mechanisms of action are associated with undesirable effects that can lead to addiction and overdose. Barbiturate-induced deaths include the high-profile suicides of the famous American actress Marilyn Monroe and Ryunosuke Akutagawa, a prolific Japanese writer and poet. Soseki Natsume attempted an overdose of sleeping pills; however, he was surrounded by many disciples, and a disciple who observed the adverse effects of the drug prevented Natsume from overdosing. This example highlights that addiction depends on the social relationships surrounding an individual.

[Allostasis and Homeostasis: Dynamic Adaptive Systems from a Neurophysiological Perspective].

Homeostasis is a well-known physiological principle that refers to a stable internal environment. Allostasis, on the other hand, is a more recently understood physiological principle based on predictive control, in which the setpoint of the internal environment is dynamically adjusted to increase control efficiency. The concept of allostasis continues to evolve with advances in the field of neuroscience. This overview introduces some new findings and broadens the concept of allostasis to apply it to the inter-regulation of the cognitive, somatic, and autonomic nervous systems. This article will describe on way in which biological systems alter themselves to adapt to their external and internal environments.

Gad1 knock-out rats exhibit abundant spike-wave discharges in EEG, exacerbated with valproate treatment.

Objective: To elucidate the functional role of gamma-aminobutyric acid (GABA)-ergic inhibition in suppressing epileptic brain activities such as spike-wave discharge (SWD), we recorded electroencephalogram (EEG) in knockout rats for Glutamate decarboxylase 1 (Gad1), which encodes one of the two GABA-synthesizing enzymes in mammals. We also examined how anti-epileptic drug valproate (VPA) acts on the SWDs present in Gad1 rats and affects GABA synthesis in the reticular thalamic nucleus (RTN), which is known to play an essential role in suppressing SWD. Methods: Chronic EEG recordings were performed in freely moving control rats and homozygous knockout Gad1 (-/-) rats. Buzzer tones (82 dB) were delivered to the rats during EEG monitoring to test whether acoustic stimulation could interrupt ongoing SWDs. VPA was administered orally to the rats, and the change in the number of SWDs was examined. The distribution of GABA in the RTN was examined immunohistochemically. Results: SWDs were abundant in EEG from Gad1 (-/-) rats as young as 2 months old. Although SWDs were universally detected in older rats irrespective of their Gad1 genotype, SWD symptom was most severe in Gad1 (-/-) rats. Acoustic stimulation readily interrupted ongoing SWDs irrespective of the Gad1 genotype, whereas SWDs were more resistant to interruption in Gad1 (-/-) rats. VPA treatment alleviated SWD symptoms in control rats, however, counterintuitively exacerbated the symptoms in Gad1 (-/-) rats. The immunohistochemistry results indicated that GABA immunoreactivity was significantly reduced in the somata of RTN neurons in Gad1 (-/-) rats but not in their axons targeting the thalamus. VPA treatment greatly increased GABA immunoreactivity in the RTN neurons of Gad1 (-/-) rats, which is likely due to the intact GAD2, another GAD isozyme, in these neurons. Discussion: Our results revealed two opposing roles of GABA in SWD generation: suppression and enhancement of SWD. To account for these contradictory roles, we propose a model in which GABA produced by GAD1 in the RTN neuronal somata is released extrasynaptically and mediates intra-RTN inhibition.

The Development of Aptamer-Coupled Microelectrode Fiber Sensors (apta-µFS) for Highly Selective Neurochemical Sensing.

The selective and sensitive sensing of neurochemicals is essential to decipher in-brain chemistry underlying brain pathophysiology. The recent development of flexible and multifunctional polymer-based fibers has been shown useful in recording and modulating neural activities, primarily electrical ones. In this study, we were able to realize fiber-based neurochemical sensing with high sensitivity and selectivity. We achieved a generalizable method to couple aptamers, a type of synthetic receptors on the carbon composites within fibers, as microsensors for highly selective neurochemical detection. Such an aptamer-coupled microelectrode fiber sensor (apta-µFS) enables simple, label-free, and sensitive dopamine (DA) detection down to 5 nM with ultrahigh specificity across major interferents. We succeeded in monitoring DA selectively within the living brain using our apta-µFS. We further showed the proof-of-concept of using microelectronic fiber-based toolsets to target neural pathways across electrical and chemical modalities. In summary, such fiber-based toolsets hold great potential to advance multimodal mechanistic understanding of brain pathophysiology.

Non-overlapping sets of neurons encode behavioral response determinants across different tasks in the posterior medial prefrontal cortex.

Higher mammals are able to simultaneously learn and perform a wide array of complex behaviors, which raises questions about how the neural representations of multiple tasks coexist within the same neural network. Do neurons play invariant roles across different tasks? Alternatively, do the same neurons play different roles in different tasks? To address these questions, we examined neuronal activity in the posterior medial prefrontal cortex of primates while they were performing two versions of arm-reaching tasks that required the selection of multiple behavioral tactics (i.e., the internal protocol of action selection), a critical requirement for the activation of this area. During the performance of these tasks, neurons in the pmPFC exhibited selective activity for the tactics, visuospatial information, action, or their combination. Surprisingly, in 82% of the tactics-selective neurons, the selective activity appeared in a particular task but not in both. Such task-specific neuronal representation appeared in 72% of the action-selective neurons. In addition, 95% of the neurons representing visuospatial information showed such activity exclusively in one task but not in both. Our findings indicate that the same neurons can play different roles across different tasks even though the tasks require common information, supporting the latter hypothesis.

[The King's Speech: Understanding Speech Act and Stuttering].

The King's Speech is a 2010 film based on historical facts of the friendship between King George VI of England, who had a stutter, and Lionel Logue, a speech therapist from Australia who treated him. The film presents a neuroscientific examination of speech and stuttering as a disorder that forms the basis of Logue's seemingly bizarre treatment and actions.

Complementary roles of primate dorsal premotor and pre-supplementary motor areas to the control of motor sequences.

We are able to temporally organize multiple movements in a purposeful manner in everyday life. Both the dorsal premotor (PMd) and pre-supplementary motor areas (pre-SMA) are known to be involved in the performance of motor sequences. However, it is unclear how each area differentially contributes to controlling multiple motor sequences. To address this issue, we recorded single-unit activity in both areas while monkeys (one male, one female) performed sixteen motor sequences. Each sequence comprised either a series of two identical movements (repetitive) or two different movements (non-repetitive). The sequence was initially instructed with visual signals but had to be remembered thereafter. Here we showed that the activity of single neurons in both areas transitioned from reactive- to predictive encoding while motor sequences were memorized. In the memory-guided trials, in particular, the activity of PMd cells preferentially represented the second movement in the sequence leading to a reward generally irrespective of the first movement. Such activity frequently began even before the first movement in a prospective manner, and was enhanced in non-repetitive sequences. Behaviorally, a lack of the activity enhancement often resulted in premature execution of the second movement. In contrast, cells in pre-SMA instantiated particular sequences of actions by coordinating switching or non-switching movements in sequence. Our findings suggest that PMd and pre-SMA play complementary roles within behavioral contexts: PMd preferentially controls the movement that leads to a reward rather than the sequence per se, whereas pre-SMA coordinates all elements in a sequence by integrating temporal orders of multiple movements.Significance statement:Although both PMd and pre-SMA are involved in the control of motor sequences, it is not clear how these two areas contribute to coordination of sequential movements differently. To address this issue, we directly compared neuronal activity in the two areas recorded while monkeys memorized and performed multiple motor sequences. Our findings suggest that PMd preferentially represents the final action that ultimately leads to a reward in a prospective manner, whereas the pre-SMA coordinates switching among multiple actions within the context of the sequence. Our findings are of significance to understand the distinct roles for motor-related areas in the planning and executing motor sequences and the pathophysiology of apraxia and/or Parkinson's diseases that disables skilled motor actions.

Reinforcement Learning Model With Dynamic State Space Tested on Target Search Tasks for Monkeys: Extension to Learning Task Events.

Learning is a crucial basis for biological systems to adapt to environments. Environments include various states or episodes, and episode-dependent learning is essential in adaptation to such complex situations. Here, we developed a model for learning a two-target search task used in primate physiological experiments. In the task, the agent is required to gaze one of the four presented light spots. Two neighboring spots are served as the correct target alternately, and the correct target pair is switched after a certain number of consecutive successes. In order for the agent to obtain rewards with a high probability, it is necessary to make decisions based on the actions and results of the previous two trials. Our previous work achieved this by using a dynamic state space. However, to learn a task that includes events such as fixation to the initial central spot, the model framework should be extended. For this purpose, here we propose a "history-in-episode architecture." Specifically, we divide states into episodes and histories, and actions are selected based on the histories within each episode. When we compared the proposed model including the dynamic state space with the conventional SARSA method in the two-target search task, the former performed close to the theoretical optimum, while the latter never achieved target-pair switch because it had to re-learn each correct target each time. The reinforcement learning model including the proposed history-in-episode architecture and dynamic state scape enables episode-dependent learning and provides a basis for highly adaptable learning systems to complex environments.

Shape and Rule Information Is Reflected in Different Local Field Potential Frequencies and Different Areas of the Primate Lateral Prefrontal Cortex.

The lateral prefrontal cortex (LFPC) plays a crucial role in executive function by adaptively storing behavior-relevant information as working memory. Neural mechanisms associated with local field potentials (LFPs) may underlie the adaptive properties of the LFPC. Here, we analyzed how LFPs recorded from the monkey LFPC are modulated by the crucial factors of a shape manipulation task. In this task, the test shape is transformed by manipulating a lever to match the size and orientation of the sample shape. The subject is required to temporarily memorize the rules such as the arm-movement-manipulation relationship and the sample shape to generate the sequential behavior of operations. In the present study, we focused on task variables about shape and rules, and examined among which aspects distinguish the ventral and dorsal sides of the LFPC. We found that the transformed shape in the sample period strongly affected the theta and delta waves in the delay period on the ventral side, while the arm-manipulation assignment influenced the gamma components on the dorsal side. These findings suggest that area- and frequency-selective LFP modulations are involved in dynamically recruiting different behavior-relevant information in the LFPC.

[Hermeneutics of Neural Activity: The Relationship Between Brain Activity from Multiple Perspectives].

From the 20th century to the 21st century, rapid progress has been made in the field of neuroscience due to the application of single-cell and multi-channel recording techniques to awake animals while they are engaged in behavioral tasks. As a result, neuroscience has challenged areas that were previously thought to fall within the field of psychology. However, several problems have been pointed out regarding the discourse and literacy of neuroscience. Therefore, in this essay, I will discuss interpretations of neural activity from three perspectives: causality, part-whole relationships, and inside-out vs. outside-in research approaches.

Non-additive activity modulation during a decision making task involving tactic selection.

Human brain imaging has revealed that stimulus-induced activity does generally not simply add to the pre-stimulus activity, but rather builds in a non-additive way on this activity. Here we investigate this subject at the single neuron level and address the question whether and to what extent a strong form of non-additivity where activity drops post-cue is present in different areas of monkey cortex, including prefrontal and agranular frontal areas, during a perceptual decision making task involving action and tactic selection. Specifically we analyze spike train data recorded in vivo from the posterior dorsomedial prefrontal cortex (pmPFC), the supplementary motor area (SMA) and the presupplementary motor area (pre-SMA). For each neuron, we compute the ratio of the trial-averaged pre-stimulus spike count to the trial-averaged post-stimulus count. We also perform the ratio and averaging procedures in reverse order. We find that the statistics of these quantities behave differently across areas. pmPFC involved in tactic selection shows stronger non-additivity compared to the two other areas which more generically just increase their firing rate pos-stimulus. pmPFC behaved more similarly to pre-SMA, a likely consequence of the reciprocal connections between these areas. The trial-averaged ratio statistic was reproduced by a surrogate inhomogeneous Poisson process in which the measured trial-averaged firing rate for a given neuron is used as its time-dependent rate. Principal component analysis (PCA) of the trial-averaged firing rates of neuronal ensembles further reveals area-specific time courses of response to the stimulus, including latency to peak neural response, for the typical population activity. Our work demonstrates subtle forms of area-specific non-additivity based on the fine variability structure of pre- and post-stimulus spiking activity on the single neuron level. It also reveals significant differences between areas for PCA and surrogate analysis, complementing previous observations of regional differences based solely on post-stimulus responses. Moreover, we observe regional differences in non-additivity which are related to the monkey's successful tactic selection and decision making. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s11571-021-09702-0.

Optogenetic stimulus-triggered acquisition of seizure resistance.

Unlike an electrical circuit, the hardware of the brain is susceptible to change. Repeated electrical brain stimulation mimics epileptogenesis. After such "kindling" process, a moderate stimulus would become sufficient in triggering a severe seizure. Here, we report that optogenetic neuronal stimulation can also convert the rat brain to a hyperexcitable state. However, continued stimulation once again converted the brain to a state that was strongly resistant to seizure induction. Histochemical examinations showed that moderate astrocyte activation was coincident with resilience acquisition. Administration of an adenosine A1 receptor antagonist instantly reverted the brain back to a hyperexcitable state, suggesting that hyperexcitability was suppressed by adenosine. Furthermore, an increase in basal adenosine was confirmed using in vivo microdialysis. Daily neuron-to-astrocyte signaling likely prompted a homeostatic increase in the endogenous actions of adenosine. Our data suggest that a certain stimulation paradigm could convert the brain circuit resilient to epilepsy without exogenous drug administration.

[Reading Dostoevsky and Polyphony from the Neuroscience Perspective].

Dostoevsky suffered from epilepsy all his life. It is known that the characters with epilepsy and the epileptic seizure held an important key in his story. While interpreting his illness and works from a neuroscience perspective, this short essay goes further and examines the structure of polyphony advocated by Bakhtin about Dostoevsky's works. Neuroscientific significance could be found in the polyphonic structure of his work consisting of the multi-layered structure of dialogue and narrative.

Heterogeneous GAD65 Expression in Subtypes of GABAergic Neurons Across Layers of the Cerebral Cortex and Hippocampus.

Gamma-aminobutyric acid (GABA), a major inhibitory transmitter in the central nervous system, is synthesized via either of two enzyme isoforms, GAD65 or GAD67. GAD65 is synthesized in the soma but functions at synaptic terminals in an activity-dependent manner, playing a distinct role in excitatory-inhibitory balance. However, the extent to which each GABAergic subtype expresses GAD65 in the resting state remains unclear. In this study, we compared GAD65 expression among six GABAergic subtypes: NPY+, nNOS+, PV+, SOM+, CR+, and CCK+. According to the results, the GABAergic subtypes were classified into two groups per region based on GAD65 expression levels: high-expression (NPY+ and nNOS+) and low-expression groups (PV+, SOM+, CR+, and CCK+) in the cerebral cortex and high-expression (NPY+, nNOS+, and CCK+) and low-expression groups (PV+, SOM+, and CR+) in the hippocampus. Moreover, these expression patterns revealed a distinct laminar distribution in the cerebral cortex and hippocampus. To investigate the extent of GAD65 transport from the soma to synaptic terminals, we examined GAD65 expression in colchicine-treated rats in which GAD65 was synthesized in the soma but not transported to terminals. We found a significant positive correlation in GAD65 expression across subtypes between colchicine-treated and control rats. In summary, each GABAergic subtype exhibits a distinct GAD65 expression pattern across layers of the cerebral cortex and hippocampus. In addition, the level of GAD65 expression in the soma can be used as a proxy for the amount of GAD65 in the cytoplasm. These findings suggest that exploration of the distinct profiles of GAD65 expression among GABAergic subtypes could clarify the roles that GABAergic subtypes play in maintaining the excitatory-inhibitory balance.

Theta Dynamics Contribute to Retrieving Motor Plans after Interruptions in the Primate Premotor Area.

To achieve a behavioral goal, we often need to maintain an internal action plan against external interruption and thereafter retrieve the action plan. We recently found that the maintenance and updating of motor plans are reflected by reciprocal changes in the beta and gamma power of the local field potential (LFP) of the primate medial motor areas. In particular, the maintenance of the immediate motor plan is supported by enhanced beta oscillations. However, it is unclear how the brain manages to maintain and retrieve the internal action plan against interruptions. Here, we show that dynamic theta changes contribute to the maintenance of the action plan. Specifically, the power of the theta frequency band (4-10 Hz) of LFPs increased before and during the interruption in the dorsal premotor areas in two monkeys. Without theta enhancement before the interruption, retrieval of the internal action plan was impaired. Theta and beta oscillations showed distinct changes depending on the behavioral context. Our results demonstrate that immediate and suspended motor plans are supported by the beta and theta oscillatory components of LFPs. Motor cortical theta oscillations may contribute to bridging motor plans across behavioral interruptions in a prospective manner.

Optical manipulation of local cerebral blood flow in the deep brain of freely moving mice.

An artificial tool for manipulating local cerebral blood flow (CBF) is necessary for understanding how CBF controls brain function. Here, we generate vascular optogenetic tools whereby smooth muscle cells and endothelial cells express optical actuators in the brain. The illumination of channelrhodopsin-2 (ChR2)-expressing mice induces a local reduction in CBF. Photoactivated adenylyl cyclase (PAC) is an optical protein that increases intracellular cyclic adenosine monophosphate (cAMP), and the illumination of PAC-expressing mice induces a local increase in CBF. We target the ventral striatum, determine the temporal kinetics of CBF change, and optimize the illumination intensity to confine the effects to the ventral striatum. We demonstrate the utility of this vascular optogenetic manipulation in freely and adaptively behaving mice and validate the task- and actuator-dependent behavioral readouts. The development of vascular optogenetic animal models will help accelerate research linking vasculature, circuits, and behavior to health and disease.

Multimodal Functional Analysis Platform: 2. Development of Si Opto-Electro Multifunctional Neural Probe with Multiple Optical Waveguides and Embedded Optical Fiber for Optogenetics.

We have developed a Si opt-electro multifunctional neural probe with multiple waveguides and embedded optical fiber for highly accurate optical stimulation. The Si opt-electro multifunctional neural probe had 16 recording sites, three optical waveguides, and metal cover for suppressing light leakage. The other opt-electro multifunctional neural probe had an optical fiber in the trench of the probe shank, which leads to fewer damages to tissues. We evaluated the electrochemical properties of the recording sites and confirmed that the neural probe had suitable characteristics for neural recording. We also demonstrated the optical stimulation to the neurons expressing ChR2 using our probe. As a result, we succeeded in multisite optical stimulation and observed that no light leakage from the optical waveguides because of the metal cover. From in vivo experiments, we successfully recorded optically modulated local field potential using the fabricated Si neural probe with optical waveguides. Moreover, we applied current source density analysis to the recorded LFPs. As a result, we confirmed that the light-induced membrane current sinks in the locally stimulated area. The Si opto-electro multifunctional neural probe is one of the most versatile tools for optogenetics.

Multimodal Functional Analysis Platform: 3. Spherical Treadmill System for Small Animals.

In the application of advanced neuroscience techniques including optogenetics to small awake animals, it is often necessary to restrict the animal's movements. A spherical treadmill is a beneficial option that enables virtual locomotion of body- or head-restrained small animals. Besides, it has a wide application range, including virtual reality experiments. This chapter describes the fundamentals of a spherical treadmill for researchers who want to start experiments with it. First, we describe the physical aspect of a spherical treadmill based on the simple mechanical analysis. Next, we explain the basics of data logging and preprocessing for behavioral analysis. We also provide simple computer programs that work for the purpose.