Neural Coding of Cell Assemblies via Spike-Timing Self-Information.

Cracking brain's neural code is of general interest. In contrast to the traditional view that enormous spike variability in resting states and stimulus-triggered responses reflects noise, here, we examine the "Neural Self-Information Theory" that the interspike-interval (ISI), or the silence-duration between 2 adjoining spikes, carries self-information that is inversely proportional to its variability-probability. Specifically, higher-probability ISIs convey minimal information because they reflect the ground state, whereas lower-probability ISIs carry more information, in the form of "positive" or "negative surprisals," signifying the excitatory or inhibitory shifts from the ground state, respectively. These surprisals serve as the quanta of information to construct temporally coordinated cell-assembly ternary codes representing real-time cognitions. Accordingly, we devised a general decoding method and unbiasedly uncovered 15 cell assemblies underlying different sleep cycles, fear-memory experiences, spatial navigation, and 5-choice serial-reaction time (5CSRT) visual-discrimination behaviors. We further revealed that robust cell-assembly codes were generated by ISI surprisals constituted of ~20% of the skewed ISI gamma-distribution tails, conforming to the "Pareto Principle" that specifies, for many events-including communication-roughly 80% of the output or consequences come from 20% of the input or causes. These results demonstrate that real-time neural coding arises from the temporal assembly of neural-clique members via silence variability-based self-information codes.

Neural Coding of Appetitive Food Experiences in the Amygdala.

Real-life experiences involve the consumption of various foods, yet it is unclear how the brain distinguishes and categorizes such food experiences. Despite the crucial roles of the basolateral amygdala (BLA) in appetitive behavior and emotion, how BLA pyramidal cells and interneurons encode food experiences has not yet been well characterized. Here we employ large-scale tetrode recording techniques to investigate the coding properties of pyramidal neurons vs. fast-spiking interneurons in the BLA as mice freely consumed a variety of foods, such as biscuits, rice, milk and water. We found that putative pyramidal cells conformed to the power-of-two-based permutation logic, as postulated by the Theory of Connectivity, to generate specific-to-general neural clique-coding patterns. Many pyramidal cells exhibited firing increases specific to a given food type, while some other pyramidal cells increased firings to various combinations of multiple foods. In contrast, fast-spiking interneurons can increase or decrease firings to given food types, and were more broadly tuned to various food experiences. We further show that a subset of pyramidal cells exhibited rapid desensitization to repeated eating of the same food, correlated with rapid behavioral habituation. Finally, we provide the intuitive visualization of BLA ensemble activation patterns using the dimensionality-reduction classification method to decode real-time appetitive stimulus identity on a moment-to-moment, single trial basis. Elucidation of the neural coding patterns in the BLA provides a key insight into how the brain's emotion and memory circuits performs the computational operation of pattern discrimination and categorization of natural food experiences.

Adult forebrain NMDA receptors gate social motivation and social memory.

Motivation to engage in social interaction is critical to ensure normal social behaviors, whereas dysregulation in social motivation can contribute to psychiatric diseases such as schizophrenia, autism, social anxiety disorders and post-traumatic stress disorder (PTSD). While dopamine is well known to regulate motivation, its downstream targets are poorly understood. Given the fact that the dopamine 1 (D1) receptors are often physically coupled with the NMDA receptors, we hypothesize that the NMDA receptor activity in the adult forebrain principal neurons are crucial not only for learning and memory, but also for the proper gating of social motivation. Here, we tested this hypothesis by examining sociability and social memory in inducible forebrain-specific NR1 knockout mice. These mice are ideal for exploring the role of the NR1 subunit in social behavior because the NR1 subunit can be selectively knocked out after the critical developmental period, in which NR1 is required for normal development. We found that the inducible deletion of the NMDA receptors prior to behavioral assays impaired, not only object and social recognition memory tests, but also resulted in profound deficits in social motivation. Mice with ablated NR1 subunits in the forebrain demonstrated significant decreases in sociability compared to their wild type counterparts. These results suggest that in addition to its crucial role in learning and memory, the NMDA receptors in the adult forebrain principal neurons gate social motivation, independent of neuronal development.

Importance of the GluN2B carboxy-terminal domain for enhancement of social memories.

The N-methyl-D-aspartate (NMDA) receptor is known to be necessary for many forms of learning and memory, including social recognition memory. Additionally, the GluN2 subunits are known to modulate multiple forms of memory, with a high GluN2A:GluN2B ratio leading to impairments in long-term memory, while a low GluN2A:GluN2B ratio enhances some forms of long-term memory. Here, we investigate the molecular motif responsible for the differences in social recognition memory and olfactory memory in the forebrain-specific transgenic GluN2A overexpression mice and the forebrain-specific transgenic GluN2B overexpression mice by using two transgenic mouse lines that overexpress chimeric GluN2 subunits. The transgenic chimeric GluN2 subunit mice were tested for their ability to learn and remember fruit scents, male juveniles of the same strain, females of the same strain, male juveniles of another strain, and rodents of another species. The data presented here demonstrate that the GluN2B carboxy-terminal domain is necessary for enhanced social recognition memory in GluN2B transgenic overexpression mice. Furthermore, the GluN2A carboxy-terminal domain is responsible for the impaired long-term olfactory and social memory observed in the GluN2A overexpression mice.

Optimization of large-scale mouse brain connectome via joint evaluation of DTI and neuron tracing data.

Tractography based on diffusion tensor imaging (DTI) data has been used as a tool by a large number of recent studies to investigate structural connectome. Despite its great success in offering unique 3D neuroanatomy information, DTI is an indirect observation with limited resolution and accuracy and its reliability is still unclear. Thus, it is essential to answer this fundamental question: how reliable is DTI tractography in constructing large-scale connectome? To answer this question, we employed neuron tracing data of 1772 experiments on the mouse brain released by the Allen Mouse Brain Connectivity Atlas (AMCA) as the ground-truth to assess the performance of DTI tractography in inferring white matter fiber pathways and inter-regional connections. For the first time in the neuroimaging field, the performance of whole brain DTI tractography in constructing a large-scale connectome has been evaluated by comparison with tracing data. Our results suggested that only with the optimized tractography parameters and the appropriate scale of brain parcellation scheme, can DTI produce relatively reliable fiber pathways and a large-scale connectome. Meanwhile, a considerable amount of errors were also identified in optimized DTI tractography results, which we believe could be potentially alleviated by efforts in developing better DTI tractography approaches. In this scenario, our framework could serve as a reliable and quantitative test bed to identify errors in tractography results which will facilitate the development of such novel tractography algorithms and the selection of optimal parameters.

On initial Brain Activity Mapping of episodic and semantic memory code in the hippocampus.

It has been widely recognized that the understanding of the brain code would require large-scale recording and decoding of brain activity patterns. In 2007 with support from Georgia Research Alliance, we have launched the Brain Decoding Project Initiative with the basic idea which is now similarly advocated by BRAIN project or Brain Activity Map proposal. As the planning of the BRAIN project is currently underway, we share our insights and lessons from our efforts in mapping real-time episodic memory traces in the hippocampus of freely behaving mice. We show that appropriate large-scale statistical methods are essential to decipher and measure real-time memory traces and neural dynamics. We also provide an example of how the carefully designed, sometime thinking-outside-the-box, behavioral paradigms can be highly instrumental to the unraveling of memory-coding cell assembly organizing principle in the hippocampus. Our observations to date have led us to conclude that the specific-to-general categorical and combinatorial feature-coding cell assembly mechanism represents an emergent property for enabling the neural networks to generate and organize not only episodic memory, but also semantic knowledge and imagination.

c-fos regulates neuronal excitability and survival.

Excitotoxicity is a process in which glutamate or other excitatory amino acids induce neuronal cell death. Accumulating evidence suggests that excitotoxicity may contribute to human neuronal cell loss caused by acute insults and chronic degeneration in the central nervous system. The immediate early gene (IEG) c-fos encodes a transcription factor. The c-Fos proteins form heterodimers with Jun family proteins, and the resulting AP-1 complexes regulate transcription by binding to the AP-1 sequence found in many cellular genes. Emerging evidence suggests that c-fos is essential in regulating neuronal cell survival versus death. Although c-fos is induced by neuronal activity, including kainic acid-induced seizures, whether and how c-fos is involved in excitotoxicity is still unknown. To address this issue, we generated a mouse in which c-fos expression is largely eliminated in the hippocampus. We found that these mutant mice have more severe kainic acid-induced seizures, increased neuronal excitability and neuronal cell death, compared with control mice. Moreover, c-Fos regulates the expression of the kainic acid receptor GluR6 and brain-derived neurotrophic factor (BDNF), both in vivo and in vitro. Our results suggest that c-fos is a genetic regulator for cellular mechanisms mediating neuronal excitability and survival.

Cognition enhancement strategies.

Many mental disorders and neurodegenerative and neurodevelopmental diseases involve cognitive deficits. Remarkable advances and new technologies are providing a clearer picture of the molecular basis of cognition. In conjunction with an SFN2010 symposium, we provided here a brief overview of the molecular mechanisms of cognition, with emphasis on the development of treatments for cognitive disorders. Activity-dependent changes in gene expression and protein synthesis integrate with synapse selection to form memory circuits. A neuronal activity-dependent molecular tagging system that uses the gene expression program to record memory circuit formation represents one new tool to study cognition. Regulation of protein translation, protein degradation, cytoskeletal dynamics, extracellular matrix interactions, second messenger signaling, and neurotransmitter receptor trafficking and function are all components of synaptic remodeling essential for cognition. Selective targeting of specific effectors in these processes, such as NMDA receptors, may serve as an effective strategy to treat cognitive deficits.

CaMKII activation state underlies synaptic labile phase of LTP and short-term memory formation.

BACKGROUND: The labile state of short-term memory has been known for more than a century. It has been frequently reported that immediate postlearning intervention can readily disrupt newly formed memories. However, the molecular and cellular mechanisms underlying the labile state of new memory are not understood. RESULTS: Using a bump-and-hole-based chemical-genetic method, we have rapidly and selectively manipulated alpha CaMKII activity levels in the mouse forebrain during various stages of the short-term memory processes. We find that a rapid shift in the alpha CaMKII activation status within the immediate 10 min after learning severely disrupts short-term memory formation. The same manipulation beyond the 15 min after learning has no effect, suggesting a critical time window for CaMKII action. We further show that during this same 10 min time window only, shifting in CaMKII activation state is capable of altering newly established synaptic weights and/or patterns. CONCLUSION: The initial 10 min of memory formation and long-term potentiation are sensitive to inducible genetic upregulation of alphaCaMKII activity. Our results suggest that molecular dynamics of CaMKII play an important role in underlying synaptic labile state and representation of short-term memory during this critical time window.

Efficient reproduction of cynomolgus monkey using pronuclear embryo transfer technique.

One of the technical bottlenecks in producing nonhuman primate models is that current assisted reproductive techniques, such as in vitro culture and frozen conservation of multicell-stage embryos, often result in poor embryo quality and subsequently lead to low birth rates. We investigated whether pronuclear embryo transfer can be used as an effective means for improving pregnancy and live birth rates of nonhuman primates. We collected 174 metaphase II oocytes by laparoscopy from 22 superovulated mature females and then fertilized these eggs using either in vitro fertilization or intracytoplasmic sperm injection, resulting in a 33.3% and a 50% fertilization rate, respectively. These 66 fertilized pronuclear-stage embryos were then tubally transferred to 30 recipients and led to 7 births and 1 abortion. Importantly, we observed that the highest live birth rate of approximately 64% was obtained when the transfer of pronuclear embryos was performed in the presence of new corpus luteum in the ovary of recipients between 24 h and 36 h after estradiol peak. Therefore, our experiments demonstrate that by matching the critical time window in the recipient's reproductive cycle for achieving optimal embryo-uterine synchrony, pronuclear embryo transfer technology can significantly improve the pregnancy rate and live birth of healthy baby monkeys. This efficient method should be valuable to the systematic efforts in construction of various transgenic primate disease models.

Functional disturbances in the striatum by region-specific ablation of NMDA receptors.

To study the role of NMDA receptors in dopamine signaling of the striatum, the brain area that receives glutamatergic inputs from various cortical areas and most dopaminergic inputs, we generated striatum-specific NMDA receptor-deficient mice. The mutant pups showed reduced food intake and retarded growth starting at the second postnatal week and died on approximately postnatal day 20 (P20). The time course of postnatal lethality is similar to that of compound mutant, double knockout of dopamine D1/D2 receptors, or genetically engineered dopamine-deficient mouse. In vivo electrophysiological recordings in the mutant pups showed that frequencies in the range of gamma oscillation were reduced in the striatal circuits. Moreover, the number of functional dopamine receptors in the striatum as measured by D1- and D2-binding experiments was greatly diminished in the mutants as compared with control animals. A consequence of diminished dopamine binding in the striatum manifested in an increase of locomotor activity. The administration of D1/D2 agonists paradoxically reduced the hyperactivity of the mutant mice as compared with an increase in locomotor activity in control mice. These results demonstrate that the NMDA receptor plays an essential role in the integration of dopamine signaling in the striatum and that is required in behavioral function.

In vivo evidence for NMDA receptor-mediated excitotoxicity in a murine genetic model of Huntington disease.

N-methyl-D-aspartate receptor (NMDAR)-mediated excitotoxicity is implicated as a proximate cause of neurodegeneration in Huntington Disease (HD). This hypothesis has not been tested rigorously in vivo. NMDAR-NR2B subunits are a major NR2 subunit expressed by striatal medium spiny neurons that degenerate in HD. To test the excitotoxic hypothesis, we crossed a well validated murine genetic model of HD (Hdh((CAG)150)) with a transgenic line overexpressing NMDAR-NR2B subunits. In the resulting double-mutant line, we show exacerbation of selective striatal neuron degeneration. This is the first direct in vivo evidence of NR2B-NMDAR-mediated excitotoxicity in the context of HD. Our results are consistent with previous suggestions that direct and/or indirect interactions of mutant huntingtin with NMDARs are a proximate cause of neurodegeneration in HD.

Neuronal PPARgamma deficiency increases susceptibility to brain damage after cerebral ischemia.

Peroxisome proliferator-activated receptor gamma (PPARgamma) plays a role in regulating a myriad of biological processes in virtually all brain cell types, including neurons. We and others have reported recently that drugs which activate PPARgamma are effective in reducing damage to brain in distinct models of brain disease, including ischemia. However, the cell type responsible for PPARgamma-mediated protection has not been established. In response to ischemia, PPARgamma gene is robustly upregulated in neurons, suggesting that neuronal PPARgamma may be a primary target for PPARgamma-agonist-mediated neuroprotection. To understand the contribution of neuronal PPARgamma to ischemic injury, we generated conditional neuron-specific PPARgamma knock-out mice (N-PPARgamma-KO). These mice are viable and appeared to be normal with respect to their gross behavior and brain anatomy. However, neuronal PPARgamma deficiency caused these mice to experience significantly more brain damage and oxidative stress in response to middle cerebral artery occlusion. The primary cortical neurons harvested from N-PPARgamma-KO mice, but not astroglia, exposed to ischemia in vitro demonstrated more damage and a reduced expression of numerous key gene products that could explain increased vulnerability, including SOD1 (superoxide dismutase 1), catalase, glutathione S-transferase, uncoupling protein-1, or transcription factor liver X receptor-alpha. Also, PPARgamma agonist-based neuroprotective effect was lost in neurons from N-PPARgamma neurons. Therefore, we conclude that PPARgamma in neurons play an essential protective function and that PPARgamma agonists may have utility in neuronal self-defense, in addition to their well established anti-inflammatory effect.

A critical role of the adenosine A2A receptor in extrastriatal neurons in modulating psychomotor activity as revealed by opposite phenotypes of striatum and forebrain A2A receptor knock-outs.

The function of striatal adenosine A(2A) receptors (A(2A)Rs) is well recognized because of their high expression levels and the documented antagonistic interaction between A(2A)Rs and dopamine D(2) receptors in the striatum. However, the role of extrastriatal A(2A)Rs in modulating psychomotor activity is largely unexplored because of the low level of expression and lack of tools to distinguish A(2A)Rs in intrinsic striatal versus nonstriatal neurons. Here, we provided direct evidence for the critical role of A(2A)Rs in extrastriatal neurons in modulating psychomotor behavior using newly developed striatum-specific A(2A)R knock-out (st-A(2A)R KO) mice in comparison with forebrain-specific A(2A)R KO (fb-A(2A)R KO) mice. In contrast to fb-A(2A)R KO (deleting A(2A)Rs in the neurons of striatum as well as cerebral cortex and hippocampus), st-A(2A)R KO mice exhibited Cre-mediated selective deletion of the A(2A)R gene, mRNA, and proteins in the neurons (but not astrocytes and microglial cells) of the striatum only. Strikingly, cocaine- and phencyclidine-induced psychomotor activities were enhanced in st-A(2A)R KO but attenuated in fb-A(2A)R KO mice. Furthermore, selective inactivation of the A(2A)Rs in extrastriatal cells by administering the A(2A)R antagonist KW6002 into st-A(2A)R KO mice attenuated cocaine effects, whereas KW6002 administration into wild-type mice enhanced cocaine effects. These results identify a critical role of A(2A)Rs in extrastriatal neurons in providing a prominent excitatory effect on psychomotor activity. These results indicate that A(2A)Rs in striatal and extrastriatal neurons exert an opposing modulation of psychostimulant effects and provide the first direct demonstration of a predominant facilitatory role of extrastriatal A(2A)Rs.

Towards transgenic primates: What can we learn from mouse genetics?

Considering the great physiological and behavioral similarities with humans, monkeys represent the ideal models not only for the study of complex cognitive behavior but also for the preclinical research and development of novel therapeutics for treating human diseases. Various powerful genetic technologies initially developed for making mouse models are being explored for generating transgenic primate models. We review the latest genetic engineering technologies and discuss the potentials and limitations for systematic production of transgenic primates.

Inducible and reversible NR1 knockout reveals crucial role of the NMDA receptor in preserving remote memories in the brain.

Long-term storage of information is a hallmark feature of the brain, yet routine turnover of synaptic receptors appears to be intrinsically paradoxical to this capability. To investigate how the brain preserves its delicate synaptic efficacies, we generated inducible and reversible knockout mice in which the NMDA receptor can be temporarily switched off in the forebrain specifically during the storage stage. Retention of 9-month contextual and cued fear memories is severely disrupted by prolonged, but not transient, loss of the NMDA receptor that occurs 6 months after initial training and at least 2 months prior to memory retrieval. Normal learning and memory function in subsequent tasks following the 9-month retention tests suggest that the observed retention deficits did not result from recall or performance impairment. Thus, our study reveals a hitherto unrecognized role of the NMDA receptor in dynamically maintaining the long-term synaptic stability of memory storage circuits in the brain.

Organizing principles of real-time memory encoding: neural clique assemblies and universal neural codes.

Recent identification of network-level coding units, termed neural cliques, in the hippocampus has enabled real-time patterns of memory traces to be mathematically described, directly visualized, and dynamically deciphered. These memory coding units are functionally organized in a categorical and hierarchical manner, suggesting that internal representations of external events in the brain is achieved not by recording exact details of those events, but rather by recreating its own selective pictures based on cognitive importance. This neural-clique-based hierarchical-extraction and parallel-binding process enables the brain to acquire not only large storage capacity but also abstraction and generalization capability. In addition, activation patterns of the neural clique assemblies can be converted to strings of binary codes that would permit universal categorizations of internal brain representations across individuals and species.

Molecular and systems mechanisms of memory consolidation and storage.

Until recently, memory consolidation and storage had been traditionally viewed as a permissive process derived from learning-activated molecular signaling cascades which include activations of the NMDA receptors, CaMKII, PKC, PKA and other kinases, new protein synthesis and CREB-mediated gene expression, and subsequent structural modifications at certain synapses. However, the time-scale of such a cascade is incompatible with the timescale of systems-level memory consolidation. Furthermore, increasing evidence suggests that synaptic proteins and structures are not stationary, but rather are highly dynamical and subjected to metabolic turnovers which would cause drift in synaptic efficacy and subsequently unstable neural circuits. Recent experiments using inducible gene- or protein-knockout techniques reveal that post-learning NMDA receptor and CaMKII reactivations are required for the systems-level consolidation of both hippocampal-dependent and hippocampal-independent memories. Furthermore, the reactivations of the NMDA receptors are also necessary for the long-term storage of old memories in the neural circuits. Therefore, the NMDA receptor reactivation-mediated synaptic reentry reinforcement (SRR) process may represent the unifying cellular mechanism in linking the consolidation and storage of long-term memories from the molecular level to the systems-level.

Histone Deacetylase Inhibitor Alleviates the Neurodegenerative Phenotypes and Histone Dysregulation in Presenilins-Deficient Mice.

Histone acetylation has been shown to play a crucial role in memory formation, and histone deacetylase (HDAC) inhibitor sodium butyrate (NaB) has been demonstrated to improve memory performance and rescue the neurodegeneration of several Alzheimer's Disease (AD) mouse models. The forebrain presenilin-1 and presenilin-2 conditional double knockout (cDKO) mice showed memory impairment, forebrain degeneration, tau hyperphosphorylation and inflammation that closely mimics AD-like phenotypes. In this article, we have investigated the effects of systemic administration of NaB on neurodegenerative phenotypes in cDKO mice. We found that chronic NaB treatment significantly restored contextual memory but did not alter cued memory in cDKO mice while such an effect was not permanent after treatment withdrawal. We further revealed that NaB treatment did not rescue reduced synaptic numbers and cortical shrinkage in cDKO mice, but significantly increased the neurogenesis in subgranular zone of dentate gyrus (DG). We also observed that tau hyperphosphorylation and inflammation related protein glial fibrillary acidic protein (GFAP) level were decreased in cDKO mice by NaB. Furthermore, GO and pathway analysis for the RNA-Seq data demonstrated that NaB treatment induced enrichment of transcripts associated with inflammation/immune processes and cytokine-cytokine receptor interactions. RT-PCR confirmed that NaB treatment inhibited the expression of inflammation related genes such as S100a9 and Ccl4 found upregulated in the brain of cDKO mice. Surprisingly, the level of brain histone acetylation in cDKO mice was dramatically increased and was decreased by the administration of NaB, which may reflect dysregulation of histone acetylation underlying memory impairment in cDKO mice. These results shed some lights on the possible molecular mechanisms of HDAC inhibitor in alleviating the neurodegenerative phenotypes of cDKO mice and provide a promising target for treating AD.