Interactions between Na?ve and Infected Macrophages Reduce Mycobacterium tuberculosis Viability

A high intracellular bacillary load of Mycobacterium tuberculosis in macrophages induces an atypical lysosomal cell death with early features of apoptosis that progress to necrosis within hours. Unlike classical apoptosis, this cell death mode does not appear to diminish M. tuberculosis viability. We previously reported that culturing heavily infected macrophages with naïve macrophages produced an antimicrobial effect, but only if naïve macrophages were added during the pre-necrotic phase of M. tuberculosis-induced cell death. In the present study we investigated the mechanism of antimicrobial activity in co-cultures, anticipating that efferocytosis of bacilli in apoptotic bodies would be required. Confocal microscopy revealed frustrated phagocytosis of M. tuberculosis-infected macrophages with no evidence that significant numbers of bacilli were transferred to the naïve macrophages. The antimicrobial effect of naïve macrophages was retained when they were separated from infected macrophages in transwells, and conditioned co-culture supernatants transferred antimicrobial activity to cultures of infected macrophages alone. Antimicrobial activity in macrophage co-cultures was abrogated when the naïve population was deficient in IL-1 receptor or when the infected population was deficient in inducible nitric oxide synthase. The participation of nitric oxide suggested a conventional antimicrobial mechanism requiring delivery of bacilli to a late endosomal compartment. Using macrophages expressing GFP-LC3 we observed the induction of autophagy specifically by a high intracellular load of M. tuberculosis. Bacilli were identified in LC3-positive compartments and LC3-positive compartments were confirmed to be acidified and LAMP1 positive. Thus, the antimicrobial effect of naïve macrophages acting on M. tuberculosis in heavily-infected macrophages is contact-independent. Interleukin-1 provides an afferent signal that induces an as yet unidentified small molecule which promotes nitric oxide-dependent antimicrobial activity against bacilli in autolysosomes of heavily infected macrophages. This cooperative, innate antimicrobial interaction may limit the maximal growth rate of M. tuberculosis prior to the expression of adaptive immunity in pulmonary tuberculosis.     

Reappraising Abstract Paintings after Exposure to Background Information

Can knowledge help viewers when they appreciate an artwork? Experts’ judgments of the aesthetic value of a painting often differ from the estimates of naïve viewers, and this phenomenon is especially pronounced in the aesthetic judgment of abstract paintings. We compared the changes in aesthetic judgments of naïve viewers while they were progressively exposed to five pieces of background information. The participants were asked to report their aesthetic judgments of a given painting after each piece of information was presented. We found that commentaries by the artist and a critic significantly increased the subjective aesthetic ratings. Does knowledge enable experts to attend to the visual features in a painting and to link it to the evaluative conventions, thus potentially causing different aesthetic judgments? To investigate whether a specific pattern of attention is essential for the knowledge-based appreciation, we tracked the eye movements of subjects while viewing a painting with a commentary by the artist and with a commentary by a critic. We observed that critics’ commentaries directed the viewers’ attention to the visual components that were highly relevant to the presented commentary. However, attention to specific features of a painting was not necessary for increasing the subjective aesthetic judgment when the artists’ commentary was presented. Our results suggest that at least two different cognitive mechanisms may be involved in knowledge- guided aesthetic judgments while viewers reappraise a painting.     

Preparation of myeloid derived suppressor cells (MDSC) from naive and pancreatic tumor-bearing mice using flow cytometry and automated magnetic activated cell sorting (AutoMACS)

MDSC are a heterogeneous population of immature macrophages, dendritic cells and granulocytes that accumulate in lymphoid organs in pathological conditions including parasitic infection, inflammation, traumatic stress, graft-versus-host disease, diabetes and cancer^1-7. In mice, MDSC express Mac-1 (CD11b) and Gr-1 (Ly6G and Ly6C) surface antigens⁷. It is important to note that MDSC are well studied in various tumor-bearing hosts where they are significantly expanded and suppress anti-tumor immune responses compared to naïve counterparts^7-10. However, depending on the pathological condition, there are different subpopulations of MDSC with distinct mechanisms and targets of suppression^11,12. Therefore, effective methods to isolate viable MDSC populations are important in elucidating their different molecular mechanisms of suppression in vitro and in vivo.Recently, the Ghansah group has reported the expansion of MDSC in a murine pancreatic cancer model. Our tumor-bearing MDSC display a loss of homeostasis and increased suppressive function compared to naïve MDSC ¹³. MDSC percentages are significantly less in lymphoid compartments of naïve vs. tumor-bearing mice. This is a major caveat, which often hinders accurate comparative analyses of these MDSC. Therefore, enriching Gr-1⁺ leukocytes from naïve mice prior to Fluorescence Activated Cell Sorting (FACS) enhances purity, viability and significantly reduces sort time. However, enrichment of Gr-1⁺ leukocytes from tumor-bearing mice is optional as these are in abundance for quick FACS sorting. Therefore, in this protocol, we describe a highly efficient method of immunophenotyping MDSC and enriching Gr-1⁺ leukocytes from spleens of naïve mice for sorting MDSC in a timely manner. Immunocompetent C57BL/6 mice are inoculated with murine Panc02 cells subcutaneously whereas naïve mice receive 1XPBS. Approximately 30 days post inoculation; spleens are harvested and processed into single-cell suspensions using a cell dissociation sieve. Splenocytes are then Red Blood Cell (RBC) lysed and an aliquot of these leukocytes are stained using fluorochrome-conjugated antibodies against Mac-1 and Gr-1 to immunophenotype MDSC percentages using Flow Cytometry. In a parallel experiment, whole leukocytes from naïve mice are stained with fluorescent-conjugated Gr-1 antibodies, incubated with PE-MicroBeads and positively selected using an automated Magnetic Activated Cell Sorting (autoMACS) Pro Separator. Next, an aliquot of Gr-1⁺ leukocytes are stained with Mac-1 antibodies to identify the increase in MDSC percentages using Flow Cytometry. Now, these Gr1⁺ enriched leukocytes are ready for FACS sorting of MDSC to be used in comparative analyses (naïve vs. tumor- bearing) in in vivo and in vitro assays.     

Immune Senescence: Relative Contributions of Age and Cytomegalovirus Infection

Immune senescence, defined as the age-associated dysregulation and dysfunction of the immune system, is characterised by impaired protective immunity and decreased efficacy of vaccines. Recent clinical, epidemiological and immunological studies suggest that Cytomegalovirus (CMV) infection may be associated with accelerated immune senescence, possibly by restricting the naïve T cell repertoire. However, direct evidence whether and how CMV-infection is implicated in immune senescence is still lacking. In this study, we have investigated whether latent mouse CMV (MCMV) infection with or without thymectomy (Tx) alters antiviral immunity of young and aged mice. After infection with lymphocytic choriomeningitis virus (LCMV) or Vaccinia virus, specific antiviral T cell responses were significantly reduced in old, old MCMV-infected and/or Tx mice compared to young mice. Importantly, control of LCMV replication was more profoundly impaired in aged MCMV-infected mice compared to age-matched MCMV-naïve or young mice. In addition, latent MCMV infection was associated with slightly reduced vaccination efficacy in old Tx mice. In contrast to the prevailing hypothesis of a CMV-mediated restriction of the naïve T cell repertoire, we found similar naïve T cell numbers in MCMV-infected and non-infected mice, whereas ageing and Tx clearly reduced the naïve T cell pool. Instead, MCMV-infection expanded the total CD8⁺ T cell pool by a massive accumulation of effector memory T cells. Based on these results, we propose a new model of increased competition between CMV-specific memory T cells and any ‘de novo’ immune response in aged individuals. In summary, our results directly demonstrate in a mouse model that latent CMV-infection impairs immunity in old age and propagates immune senescence.     

What We Observe Is Biased by What Other People Tell Us: Beliefs about the Reliability of Gaze Behavior Modulate Attentional Orienting to Gaze Cues

For effective social interactions with other people, information about the physical environment must be integrated with information about the interaction partner. In order to achieve this, processing of social information is guided by two components: a bottom-up mechanism reflexively triggered by stimulus-related information in the social scene and a top-down mechanism activated by task-related context information. In the present study, we investigated whether these components interact during attentional orienting to gaze direction. In particular, we examined whether the spatial specificity of gaze cueing is modulated by expectations about the reliability of gaze behavior. Expectations were either induced by instruction or could be derived from experience with displayed gaze behavior. Spatially specific cueing effects were observed with highly predictive gaze cues, but also when participants merely believed that actually non-predictive cues were highly predictive. Conversely, cueing effects for the whole gazed-at hemifield were observed with non-predictive gaze cues, and spatially specific cueing effects were attenuated when actually predictive gaze cues were believed to be non-predictive. This pattern indicates that (i) information about cue predictivity gained from sampling gaze behavior across social episodes can be incorporated in the attentional orienting to social cues, and that (ii) beliefs about gaze behavior modulate attentional orienting to gaze direction even when they contradict information available from social episodes.     

Phonological representations are unconsciously used when processing complex, non-speech signals

Neuroimaging studies of speech processing increasingly rely on artificial speech-like sounds whose perceptual status as speech or non-speech is assigned by simple subjective judgments; brain activation patterns are interpreted according to these status assignments. The naïve perceptual status of one such stimulus, spectrally-rotated speech (not consciously perceived as speech by naïve subjects), was evaluated in discrimination and forced identification experiments. Discrimination of variation in spectrally-rotated syllables in one group of naïve subjects was strongly related to the pattern of similarities in phonological identification of the same stimuli provided by a second, independent group of naïve subjects, suggesting either that (1) naïve rotated syllable perception involves phonetic-like processing, or (2) that perception is solely based on physical acoustic similarity, and similar sounds are provided with similar phonetic identities. Analysis of acoustic (Euclidean distances of center frequency values of formants) and phonetic similarities in the perception of the vowel portions of the rotated syllables revealed that discrimination was significantly and independently influenced by both acoustic and phonological information. We conclude that simple subjective assessments of artificial speech-like sounds can be misleading, as perception of such sounds may initially and unconsciously utilize speech-like, phonological processing.     

Perception of Social Interactions for Spatially Scrambled Biological Motion

It is vitally important for humans to detect living creatures in the environment and to analyze their behavior to facilitate action understanding and high-level social inference. The current study employed naturalistic point-light animations to examine the ability of human observers to spontaneously identify and discriminate socially interactive behaviors between two human agents. Specifically, we investigated the importance of global body form, intrinsic joint movements, extrinsic whole-body movements, and critically, the congruency between intrinsic and extrinsic motions. Motion congruency is hypothesized to be particularly important because of the constraint it imposes on naturalistic action due to the inherent causal relationship between limb movements and whole body motion. Using a free response paradigm in Experiment 1, we discovered that many naïve observers (55%) spontaneously attributed animate and/or social traits to spatially-scrambled displays of interpersonal interaction. Total stimulus motion energy was strongly correlated with the likelihood that an observer would attribute animate/social traits, as opposed to physical/mechanical traits, to the scrambled dot stimuli. In Experiment 2, we found that participants could identify interactions between spatially-scrambled displays of human dance as long as congruency was maintained between intrinsic/extrinsic movements. Violating the motion congruency constraint resulted in chance discrimination performance for the spatially-scrambled displays. Finally, Experiment 3 showed that scrambled point-light dancing animations violating this constraint were also rated as significantly less interactive than animations with congruent intrinsic/extrinsic motion. These results demonstrate the importance of intrinsic/extrinsic motion congruency for biological motion analysis, and support a theoretical framework in which early visual filters help to detect animate agents in the environment based on several fundamental constraints. Only after satisfying these basic constraints could stimuli be evaluated for high-level social content. In this way, we posit that perceptual animacy may serve as a gateway to higher-level processes that support action understanding and social inference.     

I See What You Mean: How Attentional Selection Is Shaped by Ascribing Intentions to Others

The ability to understand and predict others’ behavior is essential for successful interactions. When making predictions about what other humans will do, we treat them as intentional systems and adopt the intentional stance, i.e., refer to their mental states such as desires and intentions. In the present experiments, we investigated whether the mere belief that the observed agent is an intentional system influences basic social attention mechanisms. We presented pictures of a human and a robot face in a gaze cuing paradigm and manipulated the likelihood of adopting the intentional stance by instruction: in some conditions, participants were told that they were observing a human or a robot, in others, that they were observing a human-like mannequin or a robot whose eyes were controlled by a human. In conditions in which participants were made to believe they were observing human behavior (intentional stance likely) gaze cuing effects were significantly larger as compared to conditions when adopting the intentional stance was less likely. This effect was independent of whether a human or a robot face was presented. Therefore, we conclude that adopting the intentional stance when observing others’ behavior fundamentally influences basic mechanisms of social attention. The present results provide striking evidence that high-level cognitive processes, such as beliefs, modulate bottom-up mechanisms of attentional selection in a top-down manner.     

Cultural evolution and perpetuation of arbitrary communicative conventions in experimental microsocieties

Previous studies have shown that iconic graphical signs can evolve into symbols through repeated usage within dyads and interacting communities. Here we investigate the evolution of graphical signs over chains of participants. In these chains (or “replacement microsocieties”), membership of an interacting group changed repeatedly such that the most experienced members were continually replaced by naïve participants. Signs rapidly became symbolic, such that they were mutually incomprehensible across experienced members of different chains, and new entrants needed to learn conventionalised meanings. An objective measure of graphical complexity (perimetric complexity) showed that the signs used within the microsocieties were becoming progressively simplified over successive usage. This is the first study to show that the signs that evolve in graphical communication experiments can be transmitted to, and spontaneously adopted by, naïve participants. This provides critical support for the view that human communicative symbols could have evolved culturally from iconic representations.     

Lymphoid Tissue Damage in HIV-1 Infection Depletes Na?ve T Cells and Limits T Cell Reconstitution after Antiretroviral Therapy

Highly active antiretroviral therapy (HAART) can suppress HIV-1 replication and normalize the chronic immune activation associated with infection, but restoration of naïve CD4⁺ T cell populations is slow and usually incomplete for reasons that have yet to be determined. We tested the hypothesis that damage to the lymphoid tissue (LT) fibroblastic reticular cell (FRC) network contributes to naïve T cell loss in HIV-1 infection by restricting access to critical factors required for T cell survival. We show that collagen deposition and progressive loss of the FRC network in LTs prior to treatment restrict both access to and a major source of the survival factor interleukin-7 (IL-7). As a consequence, apoptosis within naïve T cell populations increases significantly, resulting in progressive depletion of both naïve CD4⁺ and CD8⁺ T cell populations. We further show that the extent of loss of the FRC network and collagen deposition predict the extent of restoration of the naïve T cell population after 6 month of HAART, and that restoration of FRC networks correlates with the stage of disease at which the therapy is initiated. Because restoration of the FRC network and reconstitution of naïve T cell populations are only optimal when therapy is initiated in the early/acute stage of infection, our findings strongly suggest that HAART should be initiated as soon as possible. Moreover, our findings also point to the potential use of adjunctive anti-fibrotic therapies to avert or moderate the pathological consequences of LT fibrosis, thereby improving immune reconstitution.     

In vivo expansion of naïve CD4+ CD25(high) FOXP3+ regulatory T cells in patients with colorectal carcinoma after IL-2 administration

Regulatory T cells (T_reg cells) are increased in context of malignancies and their expansion can be correlated with higher disease burden and decreased survival. Initially, interleukin 2 (IL-2) has been used as T-cell growth factor in clinical vaccination trials. In murine models, however, a role of IL-2 in development, differentiation, homeostasis, and function of T_reg cells was established. In IL-2 treated cancer patients a further T_reg-cell expansion was described, yet, the mechanism of expansion is still elusive. Here we report that functional T_reg cells of a naïve phenotype - as determined by CCR7 and CD45RA expression - are significantly expanded in colorectal cancer patients. Treatment of 15 UICC stage IV colorectal cancer patients with IL-2 in a phase I/II peptide vaccination trial further enlarges the already increased naïve T_reg-cell pool. Higher frequencies of T-cell receptor excision circles in naïve T_reg cells indicate IL-2 dependent thymic generation of naïve T_reg cells as a mechanism leading to increased frequencies of T_reg cells post IL-2 treatment in cancer patients. This finding could be confirmed in naïve murine T_reg cells after IL-2 administration. These results point to a more complex regulation of T_reg cells in context of IL-2 administration. Future strategies therefore might aim at combining IL-2 therapy with novel strategies to circumvent expansion and differentiation of naïve T_reg cells.     

When Does an Alien Become a Native Species? A Vulnerable Native Mammal Recognizes and Responds to Its Long-Term Alien Predator

The impact of alien predators on native prey populations is often attributed to prey naiveté towards a novel threat. Yet evolutionary theory predicts that alien predators cannot remain eternally novel; prey species must either become extinct or learn and adapt to the new threat. As local enemies lose their naiveté and coexistence becomes possible, an introduced species must eventually become ‘native’. But when exactly does an alien become a native species? The dingo (Canis lupus dingo) was introduced to Australia about 4000 years ago, yet its native status remains disputed. To determine whether a vulnerable native mammal (Perameles nasuta) recognizes the close relative of the dingo, the domestic dog (Canis lupus familiaris), we surveyed local residents to determine levels of bandicoot visitation to yards with and without resident dogs. Bandicoots in this area regularly emerge from bushland to forage in residential yards at night, leaving behind tell-tale deep, conical diggings in lawns and garden beds. These diggings were less likely to appear at all, and appeared less frequently and in smaller quantities in yards with dogs than in yards with either resident cats (Felis catus) or no pets. Most dogs were kept indoors at night, meaning that bandicoots were not simply chased out of the yards or killed before they could leave diggings, but rather they recognized the threat posed by dogs and avoided those yards. Native Australian mammals have had thousands of years experience with wild dingoes, which are very closely related to domestic dogs. Our study suggests that these bandicoots may no longer be naïve towards dogs. We argue that the logical criterion for determining native status of a long-term alien species must be once its native enemies are no longer naïve.     

Down-regulated NOD2 by immunosuppressants in peripheral blood cells in patients with SLE reduces the muramyl dipeptide-induced IL-10 production

Background

Pattern recognition receptors (PRRs) such as Toll-like receptors are aberrantly expressed of peripheral blood mononuclear cells (PBMCs) in systemic lupus erythematosus (SLE) patients, for playing immunopathological roles.

Methodology/Principal Findings

We investigated the expression and function of the PRR nucleotide-binding oligomerization domain (NOD2) in SLE. NOD2 expression in T, B lymphocytes, monocytes, myeloid dendritic cells (mDCs) and plasmacytoid dendritic cells (pDCs) was assessed in SLE patients and healthy controls (HCs) using flow cytometric analysis. Ex vivo production of cytokines from PBMCs upon NOD2 agonist muramyl dipeptide (MDP) stimulation was assessed using Cytometric Bead Array. Over-expression of NOD2 in monocytes was observed in immunosuppressant naïve SLE patients, and was positively associated with longer disease duration. Immunosuppressive therapy was an independent explanatory variable for downregulating NOD2 expression in CD8+ T, monocytes, mDCs and pDCs. Ex vivo basal productions of cytokines (IL-6, IL-8 and IL-10) were significantly increased in immunosuppressant naïve patients and patients with active disease despite immunosuppressants compared with HCs. Upon MDP stimulaiton, relative induction (%) of cytokines (IL-1β) from PBMC was significantly increased in immunosuppressant naïve patients with inactive disease, and patients with active disease despite immunosuppressant treatment compared with HCs. Immunosuppressant usage was associated with a decreased basal production and MDP induced relative induction (%) of IL-10 in patients with inactive disease compared with immunosuppressant naïve patients and HCs.

Conclusions/Significance

Bacterial exposure may increase the NOD2 expression in monocytes in immunosuppressant naïve SLE patients which can subsequently lead to aberrant activation of PBMCs to produce proinflammatory cytokines, implicating the innate immune response for extracellular pathogens in the immunopathological mechanisms in SLE. Immunosuppressant therapy may downregulate NOD2 expression in CD8+ T lymphocytes, monocytes, and DCs in SLE patients which subsequently IL-10 reduction, contributing towards the regulation of immunopathological mechanisms of SLE, at the expense of increasing risk of bacterial infection.     

The anti-CD74 humanized monoclonal antibody, milatuzumab, which targets the invariant chain of MHC II complexes, alters B-cell proliferation, migration, and adhesion molecule expression

Introduction

Targeting CD74 as the invariant chain of major histocompatibility complexes (MHC) became possible by the availability of a specific humanized monoclonal antibody, milatuzumab, which is under investigation in patients with hematological neoplasms. CD74 has been reported to regulate chemo-attractant migration of macrophages and dendritic cells, while the role of CD74 on peripheral naïve and memory B cells also expressing CD74 remains unknown. Therefore, the current study addressed the influence of milatuzumab on B-cell proliferation, chemo-attractant migration, and adhesion molecule expression.

Methods

Surface expression of CD74 on CD27^- naïve and CD27⁺ memory B cells as well as other peripheral blood mononuclear cells (PBMCs) obtained from normals, including the co-expression of CD44, CXCR4, and the adhesion molecules CD62L, β7-integrin, β1-integrin and CD9 were studied after binding of milatuzumab using multicolor flow cytometry. The influence of the antibody on B-cell proliferation and migration was analyzed in vitro in detail.

Results

In addition to monocytes, milatuzumab also specifically bound to human peripheral B cells, with a higher intensity on CD27⁺ memory versus CD27^- naïve B cells. The antibody reduced B-cell proliferation significantly but moderately, induced enhanced spontaneous and CXCL12-dependent migration together with changes in the expression of adhesion molecules, CD44, β7-integrin and CD62L, mainly of CD27^- naïve B cells. This was independent of macrophage migration-inhibitory factor as a ligand of CD74/CD44 complexes.

Conclusions

Milatuzumab leads to modestly reduced proliferation, alterations in migration, and adhesion molecule expression preferentially of CD27^- naïve B cells. It thus may be a candidate antibody for the autoimmune disease therapy by modifying B cell functions.     

Social Models Provide a Norm of Appropriate Food Intake for Young Women

It is often assumed that social models influence people’s eating behavior by providing a norm of appropriate food intake, but this hypothesis has not been directly tested. In three experiments, female participants were exposed to a low-intake model, a high-intake model, or no model (control condition). Experiments 1 and 2 used a remote-confederate manipulation and were conducted in the context of a cookie taste test. Experiment 3 used a live confederate and was conducted in the context of a task during which participants were given incidental access to food. Participants also rated the extent to which their food intake was influenced by a variety of factors (e.g., hunger, taste, how much others ate). In all three experiments, participants in the low-intake conditions ate less than did participants in the high-intake conditions, and also reported a lower perceived norm of appropriate intake. Furthermore, perceived norms of appropriate intake mediated the effects of the social model on participants’ food intake. Despite the observed effects of the social models, participants were much more likely to indicate that their food intake was influenced by taste and hunger than by the behavior of the social models. Thus, social models appear to influence food intake by providing a norm of appropriate eating behavior, but people may be unaware of the influence of a social model on their behavior.     

TSC1/2 Signaling Complex Is Essential for Peripheral Na?ve CD8+ T Cell Survival and Homeostasis in Mice

The PI3K-Akt-mTOR pathway plays crucial roles in regulating both innate and adaptive immunity. However, the role of TSC1, a critical negative regulator of mTOR, in peripheral T cell homeostasis remains elusive. With T cell-specific Tsc1 conditional knockout (Tsc1 KO) mice, we found that peripheral naïve CD8⁺ T cells but not CD4⁺ T cells were severely reduced. Tsc1 KO naïve CD8⁺ T cells showed profound survival defect in an adoptive transfer model and in culture with either stimulation of IL-7 or IL-15, despite comparable CD122 and CD127 expression between control and KO CD8⁺ T cells. IL-7 stimulated phosphorylation of Akt(S473) was diminished in Tsc1 KO naïve CD8⁺T cells due to hyperactive mTOR-mediated feedback suppression on PI3K-AKT signaling. Furthermore, impaired Foxo1/Foxo3a phosphorylation and increased pro-apoptotic Bim expression in Tsc1 KO naïve CD8⁺T cells were observed upon stimulation of IL-7. Collectively, our study suggests that TSC1 plays an essential role in regulating peripheral naïve CD8⁺ T cell homeostasis, possible via an mTOR-Akt-FoxO-Bim signaling pathway.     

Genome Reshuffling for Advanced Intercross Permutation (GRAIP): simulation and permutation for advanced intercross population analysis

Background

Advanced intercross lines (AIL) are segregating populations created using a multi-generation breeding protocol for fine mapping complex trait loci (QTL) in mice and other organisms. Applying QTL mapping methods for intercross and backcross populations, often followed by naïve permutation of individuals and phenotypes, does not account for the effect of AIL family structure in which final generations have been expanded and leads to inappropriately low significance thresholds. The critical problem with naïve mapping approaches in AIL populations is that the individual is not an exchangeable unit.

Methodology/Principal Findings

The effect of family structure has immediate implications for the optimal AIL creation (many crosses, few animals per cross, and population expansion before the final generation) and we discuss these and the utility of AIL populations for QTL fine mapping. We also describe Genome Reshuffling for Advanced Intercross Permutation, (GRAIP) a method for analyzing AIL data that accounts for family structure. GRAIP permutes a more interchangeable unit in the final generation crosses – the parental genome – and simulating regeneration of a permuted AIL population based on exchanged parental identities. GRAIP determines appropriate genome-wide significance thresholds and locus-specific P-values for AILs and other populations with similar family structures. We contrast GRAIP with naïve permutation using a large densely genotyped mouse AIL population (1333 individuals from 32 crosses). A naïve permutation using coat color as a model phenotype demonstrates high false-positive locus identification and uncertain significance levels, which are corrected using GRAIP. GRAIP also detects an established hippocampus weight locus and a new locus, Hipp9a.

Conclusions and Significance

GRAIP determines appropriate genome-wide significance thresholds and locus-specific P-values for AILs and other populations with similar family structures. The effect of family structure has immediate implications for the optimal AIL creation and we discuss these and the utility of AIL populations.     

Rapid antagonistic coevolution between strains of the social amoeba Dictyostelium discoideum

Social groups face a fundamental problem of overcoming selfish individuals capable of destroying cooperation. In the social amoeba Dictyostelium discoideum, there is evidence that some clones (‘cheaters’) contribute disproportionately to the viable spores in a fruiting body while avoiding the dead stalk cell fate. It remains unclear, however, whether this cheating is actually the product of selection. Here, I report the results of an experimental evolution study designed to test whether clones of D. discoideum will evolve resistance to cheating in the laboratory with genetic variation created only through spontaneous mutation. Two strains, one green fluorescent protein (GFP)-labelled and one wild-type, were allowed to grow and develop together before the wild-type strain was removed and replaced with a naïve strain evolving in parallel. Over the course of 10 social generations, the GFP-labelled strain reliably increased its representation in the spores relative to control populations that had never experienced the competitor. This competitive advantage extended to the non-social, vegetative growth portion of the life cycle, but not to pairwise competition with two other strains. These results indicate strong antagonism between strains, mediated by ample mutational variation for cheating and also suggest that arms races between strains in the wild may be common.     

Social transmission of fear in rats: the role of 22-kHz ultrasonic distress vocalization

Background

Social alarm calls alert animals to potential danger and thereby promote group survival. Adult laboratory rats in distress emit 22-kHz ultrasonic vocalization (USV) calls, but the question of whether these USV calls directly elicit defensive behavior in conspecifics is unresolved.

Methodology/Principal Findings

The present study investigated, in pair-housed male rats, whether and how the conditioned fear-induced 22-kHz USVs emitted by the ‘sender’ animal affect the behavior of its partner, the ‘receiver’ animal, when both are placed together in a novel chamber. The sender rats’ conditioned fear responses evoked significant freezing (an overt evidence of fear) in receiver rats that had previously experienced an aversive event but not in naïve receiver rats. Permanent lesions and reversible inactivations of the medial geniculate nucleus (MGN) of the thalamus effectively blocked the receivers’ freeezing response to the senders'' conditioned fear responses, and this occurred in absence of lesions/inactivations impeding the receiver animals'' ability to freeze and emit 22-kHz USVs to the aversive event per se.

Conclusions/Significance

These results—that prior experience of fear and intact auditory system are required for receiver rats to respond to their conspecifics'' conditioned fear responses—indicate that the 22-kHz USV is the main factor for social transmission of fear and that learning plays a crucial role in the development of social signaling of danger by USVs.     

Characterization of the Psychological,Physiological and EEG Profile of Acute Betel Quid Intoxication in Na?ve Subjects

Betel quid use and abuse is wide spread in Asia but the physiological basis of intoxication and addiction are unknown. In subjects naïve to the habit of betel quid intoxication, the psychological and physiological profile of intoxication has never been reported. We compared the effect of chewing gum or chewing betel quid, and subsequent betel quid intoxication, on psychological assessment, prospective time interval estimation, numerical and character digit span, computerized 2 choice tests and mental tasks such as reading and mathematics with concurrent monitoring of ECG, EEG and face temperature in healthy, non-sleep deprived, male subjects naïve to the habit of chewing betel quid. Betel quid intoxication, dose dependently induced tachycardia (max 30 bpm) and elevated face temperature (0.7°C) (P<0.001) above the effects observed in response to chewing gum (max 12 bpm and 0.3°C) in 12 subjects. Gross behavioral indices of working memory such as numerical or character digit span in 8 subjects, or simple visual-motor performance such as reaction speed or accuracy in a two choice scenario in 8 subjects were not affected by betel quid intoxication. Betel quid intoxication strongly influenced the psychological aspects of perception such as slowing of the prospective perception of passage of a 1 minute time interval in 8 subjects (P<0.05) and perceived increased arousal (P<0.01) and perceived decreased ability to think (P<0.05) in 31 subjects. The EEG spectral profile recorded from mental states associated with open and closed eyes, and mental tasks such as reading and eyes closed mental arithmetic were significantly modified (P<0.05) relative to chewing gum by betel quid intoxication in 10 subjects. The prevalence of betel quid consumption across a range of social and work settings warrants greater investigation of this widespread but largely under researched drug.