Welcome to the IBRO 2023 Interactive Programme

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In the presence of danger, emotional information related to the detection of threat must be encoded simultaneously with information devoted to discriminate among threats and select defensive responses. Using classical behavioral paradigms it has been shown that the activity of multiple brain regions plays a role in the expression of defensive responses in the presence of a threat. However, whether the investigated neuronal activity was associated to the elicited defensive state or was encoding information specific to each threatening situation remained to be elucidated. Among the brain regions involved in defensive responses, the prefrontal cortex is a higher-order structure known to control threat-related behaviors, yet it is unknown whether it encodes a global defensive state and/or the identity of specific threatening encounters. We used a combination of electrophysiological recordings, calcium imaging, neuronal decoding approaches and optogenetic manipulations in a novel behavioral paradigm allowing the simultaneous evaluation of distinct defensive behaviors in response to different threatening situations. Our results indicate that the dorsomedial prefrontal cortex (dmPFC) encodes a general representation of fear elicited by all threatening conditions while simultaneously encoding a specific neuronal representation of each threatening situation. Importantly, we demonstrated the persistence of the global representation of danger in error trials that instead lacked specific threat-identity representation. Consistently, the optogenetic inhibition of prefrontal neurons impaired the overall behavioral performance and the discrimination among different threats instead of promoting the expression of active or passive danger-coping strategies. Together this data indicates that the prefrontal cortex encodes both a global representation of danger and information necessary to discriminate among threats to select specific defensive behaviors.

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Stress- and anxiety- based disorders are characterized by generalized fear to stimuli that were never paired with an aversive outcome. To better understand the effects of stress on cue processing, I will first show how chronic stress impacts explicit safety-cue learning and its accompanying neural activity in the prefrontal cortex, as well as its communication with the amygdala. I will then discuss how these changes play into fear generalization where we think about discrimination of a non-threatening cue vs stronger learning of a threatening cue as separate processes.

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Our brains integrate sensory information with prior knowledge to guide our decisions and generate complex behavioural outcomes. However, through which neural pathways prior experience can adapt behavioural responses to sensory input is still unclear. The ventral lateral geniculated nucleus (vLGN), an inhibitory nucleus in the prethalamus, has recently been shown to have strong control over fear responses to innate visual threat. In addition, it has been shown that vLGN activity is modulated by prior experience. We found that vLGN receives prominent layer 5 input from cortical areas. We tested the hypothesis that inputs from the neocortex to vLGN -convey acquired knowledge about threatening visual stimuli, enabling this pathway to suppress fear responses to a potential threat when animals learn that it poses no danger.

We used a paradigm in which mice escape from an innately threatening visual stimulus. After frequently being exposed to this threat stimulus mice habituate – they stop escaping once they have learned that the stimulus does not pose a real danger. We found that silencing lateral higher visual areas (HVA) or specifically silencing their projections in vLGN during the habituation protocol prevented habituation, as mice kept escaping to the looming stimulus afterwards. In contrast, silencing visual cortical areas after habituation had no effect on escape responses. Additionally, silencing inhibitory vLGN cells in habituated mice could override the learnt behaviour, as mice resumed escaping to looming stimuli. These findings suggest that input from higher visual cortices specifically during habituation induces learning in vLGN circuits to induce suppression of fear responses. We found a subset of vLGN neurons that increased their activity after mice had habituated, these cells received direct input from the HVA. Positioning HVA with an instructive role for learning not to fear innate threatening visual stimulus.

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I will present recent work identifying two mechanisms of single neurons in the human amygdala that underlie differences in exploration rates between positive (reward-based) and negative (punishment-based) learning. Next, I will present findings about representations of valence (positive and negative) in the primate amygdala, anterior-cingulate-cortex, and substantia-innominata, and examine the interactions between these regions and further suggest novel ways to modulate these representations via a closed-loop brain-computer-interface techniques. These findings shed light on representations of valence-based learning in primate networks and suggest new ways to manipulate them.

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Several studies have been conducted to explore the interaction between the nervous system and the immune system since the 19th century. In the last decades, the autonomic nervous system has been the focus of these studies involving the modulation of inflammatory responses. The cardiovascular reflexes are physiological mechanisms that control both branches of the autonomic nervous system (i.e., sympathetic and parasympathetic) to maintain homeostasis. The arterial baroreflex activation inhibits the sympathetic outflow and increases the parasympathetic drive to control arterial pressure. On the other hand, the chemoreflex maintains cardiorespiratory homeostasis in response to changes in blood gas concentrations by activating both branches of the autonomic nervous system. The physiological activation of the cardiovascular reflexes is a useful approach to mimic the genuine role of the autonomic nervous system during inflammation. This lecture will briefly summarize our group’s recent discoveries suggesting that physiological reflex activation of the sympathetic circuit, involving the baroreflex and chemoreflex mechanisms, could modulate the cytokine response in endotoxemic rats. Although more studies are essential to show the complete pathway involved in this process, it is clear that a new mechanism is emerging for a better understanding of the interaction between the nervous system and the immune system and to improve the therapeutic strategies for treating inflammatory diseases.

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The nervous system communicates with and strongly influences the immune system. A powerful neural reflex, the inflammatory reflex, is endogenously activated when the body faces an immune challenge. Its function is to inhibit the innate immune response by preventing an excessive inflammatory reaction to bacterial or viral infections. The efferent pathway of the inflammatory reflex, termed the splanchnic anti-inflammatory pathway, comprises sympathetic fibers part of the greater splanchnic nerves, but not other sympathetic nerves, nor the vagus. This sympathetic pathway drives a coordinated anti-inflammatory response: it suppresses the release of pro-inflammatory cytokines, like tumor necrosis factor-α (TNF), while enhancing the production of the anti-inflammatory cytokine interleukin-10 (IL-10). Within the splanchnic nerves, divergent pathways control these two cytokine responses: neurally driven adrenaline, acting via β2 adrenoreceptors, regulates IL-10, while TNF is restrained by sympathetic nerves to abdominal organs, where non-β2 adrenoreceptor-mechanisms are also involved. The neural influence on immunity mediated by the splanchnic sympathetic nerves is powerful and not limited to the control of the inflammatory response. Animals with the splanchnic anti-inflammatory pathway compromised show a disinhibited innate immune response to a systemic E.coli infection that allows them to better fight and resolve the bacteremia. The comprehension of the mechanisms by which the inflammatory reflex inhibits the immune system could change the way we deal with different maladies and syndromes: on one side, we could enhance the ability of the immune system to fight an infection, while, on the other, we could inhibit the inflammatory response when exaggerated and harmful.

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The intestinal mucosa is home to an immune cell network that delicately needs to balance responses to microbiota in the gut lumen. In Inflammatory Bowel Diseases (IBD), a chronic inflammation of the intestinal mucosa (colitis) evolves and immune responses viciously turn against common microbiota for unknown reason. Mucosal damage is a key feature of IBD and healing of the mucosa is an endpoint of IBD treatment that is often difficult to achieve. Both branches of the autonomic nervous system, (parasympathetic and sympathetic) innervate the intestinal crypt and mucosal immune cells. Neuronal innervation affects mucosal immunity but also can mediate intestinal epithelial cell growth and intervention thereof could therefore serve as a potential therapeutic option to improve mucosal healing. I will discuss this as a potential original approach for IBD treatment based on recent recognition of the nervous system as an integrated regulator of inflammatory processes. The aim is to reduce chronic colitis by the targeted intervention in neuronal signaling to the intestinal mucosa and the spleen. This is timely because one can now make use of unique neural implantation devices that are developed for non-invasive use. We identified sympathetic nerves that are surprisingly potent in regulating severity of intestinal inflammation, leading us to apply specific neuronal stimulation in colitis models and human systems. Such interventions are currently tested in clinical trial in Rheumatoid Arthritis. I will discuss the route to therapy, and the hurdles thereto, using this new medicine free approach for chronic immune diseases.

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The nervous system and the immune system are two integrative systems that work collaboratively to detect threats, provide host defence, restore homeostasis, and their crosstalk is crucial for maintaining the health and well-being of the host. Recent discoveries show that neuro-immune interactions that control inflammation are an ancient component of animal physiology, found in the least complex nematodes and similarly described in higher vertebrates. Immune cells are equipped to respond to neuronal signals by expressing receptors for neuronal cell-derived molecules and, reciprocally, neurons express receptors for immune-derived cytokines and neurotransmitters, which can affect neuronal function. As such, neuro-immune interactions have been shown to be involved in multiple aspects of tissue physiology during homeostasis and pathologies, including haematopoiesis, sepsis, various chronic inflammatory disorders and even stroke. In this presentation, I will outline our group's recent work on the effect of stroke on the peripheral immune system, and describe how a distal injury in the brain can elicit diverse immune cell functional impairments and mucosal barrier breakdown. Detail mapping of the interplay between the nervous and immune systems anatomically and functionally during inflammation will likely reveal important information about the intricate network processes shared between two elaborate, highly co-evolved systems of defence and memory. Using this knowledge, it is possible to target neural pathways with pharmacological agents to resolve inflammation and provide therapeutic benefit.

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In developing brains, activity-dependent remodeling facilitates the formation of precise neuronal connectivity. Synaptic competition is known to facilitate synapse elimination; however, it has remained unknown how different synapses compete to each other within a postsynaptic cell. Here we investigate how a mitral cell in the olfactory bulb prunes all but one primary dendrite during the developmental remodeling process. We find that spontaneous activity generated within the olfactory bulb is essential. We show that strong glutamatergic inputs to one dendrite trigger branch-specific changes in RhoA activity to facilitate the pruning of the remaining dendrites: NMDAR-dependent local signals suppress RhoA to protect it from pruning; however, the subsequent neuronal depolarization induces neuron-wide activation of RhoA to prune non-protected dendrites. NMDAR-RhoA signals are also essential for the synaptic competition in the barrel cortex. Our results demonstrate a general principle whereby activity-dependent lateral inhibition across synapses establishes a discrete receptive field of a neuron.

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Why do developing brains become active before they can receive sensory input? To answer this question, we first need to find the source of the activity. We discovered that there is patterned, stimulus-independent neural activity (PSINA, ‘see-nah’) in the developing Drosophila brain. This approachably complex model system offers a track complementary to the on-going efforts to understand how developmental activity arises in the mammalian brain. PSINA engages the fly brain in globally coordinated, stereotyped cycles of active and silent phases during the last two days of metamorphosis. More recently, we found that a small population of some 2,000 neurons expressing the cation channel Trpγ are critical to PSINA: Silencing these Trpγ+ neurons attenuates activity by >90% and alters synapse development in a cell-type-specific manner. To ask if there is a minimal set of PSINA-active Trpγ+ neurons, we are functionally fractionating the population using SpaRCLIn, a new Drosophila toolkit which partitions GAL4 expression domains into constituent neuroblast lineages. To complement this on-going work and to extend our handle on PSINA beyond Trpγ, we carried out a neuronal silencing screen through neuropeptide gene expression domains. This effort revealed that manipulating neuronal activity in the expression domains of neuropeptides ITP (Ion Transport Peptide), nplp1 (Neuropeptide-like Precursor 1), Proc (Proctolin), and sNPF (Short Neuropeptide F) and their known or putative receptors significantly alters PSINA. We are now asking whether these new domains share common activity with Trpγ+ neurons or capture a different feature of a putative PSINA cascade. We expect that these studies will identify distinct cells and circuits which trigger PSINA and coordinate the activity across the developing brain. With these cells at hand, we will be able to ask how the stereotyped patterns of PSINA affect synapse and circuit assembly in the late pupa and behavior in the adult.

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Down Syndrome (DS) is the most common genetic disorder due to the abnormal triplication of chromosome 21 (trisomy21) resulting in a variety of pathological conditions of DS subjects. Among these, individuals affected by DS show with ageing the accumulation of oxidative damage associated with defects of the proteostasis network (1). DS is currently considered a human genetic model of early onset Alzheimer disease (AD). The talk will discuss the role of trisomic genes which, directly and indirectly, contribute to the occurrence of an aberrant redox-phenotype and how it contributes to the dysfunction of several cellular functions (2). Among these, we hypothesize that redox dysregulation is closely linked to metabolic defects, including reduced glucose metabolism, energy production and aberrant insulin signaling. Alteration of energy metabolism significantly contributes to accelerate the onset of Alzheimer like neruodegeneration, including amyloid and Tau neuropathology, in DS individuals.

References:

1) Barone E, Head E, Butterfield DA, Perluigi M. HNE-modified proteins in Down syndrome: Involvement in development of Alzheimer disease neuropathology. FRBM 2017;111:262-269. doi:

2) Insulin resistance, oxidative stress and mitochondrial defects in Ts65dn mice brain: A harmful synergistic path in down syndrome. Lanzillotta C et al. FRBM. doi: 10.1016/j.freeradbiomed.2021.01.042.