The postdoc position is focused on the development of BrainSTEM, a web application designed as an electronic lab notebook for describing neurophysiological experiments as well as a data-sharing platform for the community. The role involves the design of a standard language for describing experimental neuroscience, semantic search functionality, stronger adoption of the FAIR principles, and stimulating and supporting community uptake. The project is primarily funded by the NIH, through the Brain Initiative U19 Oxytocin grant. The project will include occasional travels, e.g., to New York (NYU), Brain Initiate meetings, SfN, FENS, and to pilot user labs.

Cardiac vagal motor drive originates in the brainstem's cardiac vagal motor neurons (CVNs). Despite well-established cardioinhibitory functions in health, our understanding of CVNs in disease is limited. There is a clear connection of cardiovascular regulation with metabolic and energy expenditure systems. Using high fat diet as a model, this talk will explore how metabolic dysfunction impacts the regulation of cardiac tissue through robust inhibition of CVNs. Specifically, it will present an often overlooked modality of inhibition, tonic gamma-aminobuytric acid (GABA) A-type neurotransmission using an array of techniques from single cell patch clamp electrophysiology to transgenic in vivo whole animal physiology. It also will highlight a unique interaction with the delta isoform of protein kinase C to facilitate GABA A-type receptor expression.

The primary visual cortex (V1) directly projects to the superior colliculus (SC) and is believed to provide sensory drive for eye movements. Consistent with this, a majority of saccade-related SC neurons also exhibit short-latency, stimulus-driven visual responses, which are additionally feature-tuned. However, direct neurophysiological comparisons of the visual response properties of the two anatomically-connected brain areas are surprisingly lacking, especially with respect to active looking behaviors. I will describe a series of experiments characterizing visual response properties in primate V1 and SC neurons, exploring feature dimensions like visual field location, spatial frequency, orientation, contrast, and luminance polarity. The results suggest a substantial, qualitative reformatting of SC visual responses when compared to V1. For example, SC visual response latencies are actively delayed, independent of individual neuron tuning preferences, as a function of increasing spatial frequency, and this phenomenon is directly correlated with saccadic reaction times. Such “coarse-to-fine” rank ordering of SC visual response latencies as a function of spatial frequency is much weaker in V1, suggesting a dissociation of V1 responses from saccade timing. Consistent with this, when we next explored trial-by-trial correlations of individual neurons’ visual response strengths and visual response latencies with saccadic reaction times, we found that most SC neurons exhibited, on a trial-by-trial basis, stronger and earlier visual responses for faster saccadic reaction times. Moreover, these correlations were substantially higher for visual-motor neurons in the intermediate and deep layers than for more superficial visual-only neurons. No such correlations existed systematically in V1. Thus, visual responses in SC and V1 serve fundamentally different roles in active vision: V1 jumpstarts sensing and image analysis, but SC jumpstarts moving. I will finish by demonstrating, using V1 reversible inactivation, that, despite reformatting of signals from V1 to the brainstem, V1 is still a necessary gateway for visually-driven oculomotor responses to occur, even for the most reflexive of eye movement phenomena. This is a fundamental difference from rodent studies demonstrating clear V1-independent processing in afferent visual pathways bypassing the geniculostriate one, and it demonstrates the importance of multi-species comparisons in the study of oculomotor control.

To maintain a stable and clear image of the world, our eyes reflexively follow the direction in which a visual scene is moving. Such gaze stabilization mechanisms reduce image blur as we move in the environment. In non-primate mammals, this behavior is initiated by ON-type direction-selective ganglion cells (ON-DSGCs), which detect the direction of image motion and transmit signals to brainstem nuclei that drive compensatory eye movements. However, ON-DSGCs have not yet been functionally identified in primates, raising the possibility that the visual inputs that drive this behavior instead arise in the cortex. In this talk, I will present molecular, morphological and functional evidence for identification of an ON-DSGC in macaque retina. The presence of ON-DSGCs highlights the need to examine the contribution of subcortical retinal mechanisms to normal and aberrant gaze stabilization in the developing and mature visual system. More generally, our findings demonstrate the power of a multimodal approach to study sparsely represented primate RGC types.

The taste system detects 5 major categories of ethologically relevant stimuli (sweet, bitter, umami, sour and salt) and accordingly elicits acceptance or avoidance responses. While these taste responses are innate, the taste system retains a remarkable flexibility in response to changing external and internal contexts. Taste chemicals are first recognized by dedicated taste receptor cells (TRCs) and then transmitted to the cortex via a multi-station relay. I reasoned that if I could identify taste neural substrates along this pathway, it would provide an entry to decipher how taste signals are encoded to drive innate response and modulated to facilitate adaptive response. Given the innate nature of taste responses, these neural substrates should be genetically identifiable. I therefore exploited single-cell RNA sequencing to isolate molecular markers defining taste qualities in the taste ganglion and the nucleus of the solitary tract (NST) in the brainstem, the two stations transmitting taste signals from TRCs to the brain. How taste information propagates from the ganglion to the brain is highly debated (i.e., does taste information travel in labeled-lines?). Leveraging these genetic handles, I demonstrated one-to-one correspondence between ganglion and NST neurons coding for the same taste. Importantly, inactivating one ‘line’ did not affect responses to any other taste stimuli. These results clearly showed that taste information is transmitted to the brain via labeled lines. But are these labeled lines aptly adapted to the internal state and external environment? I studied the modulation of taste signals by conflicting taste qualities in the concurrence of sweet and bitter to understand how adaptive taste responses emerge from hardwired taste circuits. Using functional imaging, anatomical tracing and circuit mapping, I found that bitter signals suppress sweet signals in the NST via top-down modulation by taste cortex and amygdala of NST taste signals. While the bitter cortical field provides direct feedback onto the NST to amplify incoming bitter signals, it exerts negative feedback via amygdala onto the incoming sweet signal in the NST. By manipulating this feedback circuit, I showed that this top-down control is functionally required for bitter evoked suppression of sweet taste. These results illustrate how the taste system uses dedicated feedback lines to finely regulate innate behavioral responses and may have implications for the context-dependent modulation of hardwired circuits in general.

How the brain creates a painful experience remains a mystery. Solving this mystery is crucial to understanding the fundamental biological processes that underlie the perception of body integrity, and to creating better, non-addictive pain treatments. My laboratory’s goal is to resolve the neural basis of pain. We aim to understand the mechanisms by which our nervous system produces and assembles the sensory-discriminative, affective-motivational, and cognitive-evaluative dimensions of pain to create this unique and critically important experience. To capture every component of the pain experience, we examine the entirety of the pain circuitry, from sensory and spinal ascending pathways to cortical/subcortical circuits and brainstem descending pain modulation systems, at the molecular, cellular, circuit and whole-animal levels. For these studies, we have invented novel behavioral paradigms to interrogate the affective and cognitive dimensions of pain in mice while simultaneously imaging and manipulating nociceptive circuits. My laboratory also investigates how opioids suppress pain. Remarkably, despite their medical and societal significance, how opium poppy alkaloids such as morphine produce profound analgesia remains largely unexplained. By identifying where and how opioids act in neural circuits, we not only establish the mechanisms of action of one of the oldest drugs known to humans, but also reveal the critical elements of the pain circuitry for developing of novel analgesics and bringing an end to the opioid epidemic.

The ability to experience pain is old in evolutionary terms. It is an experience shared across species. Acute pain is the body’s alarm system, and as such it is a good thing. Pain that persists beyond normal tissue healing time (3-4 months) is defined as chronic – it is the system gone wrong and it is not a good thing. Chronic pain has recently been classified as both a symptom and disease in its own right. It is one of the largest medical health problems worldwide with one in five adults diagnosed with the condition. The brain is key to the experience of pain and pain relief. This is the place where pain emerges as a perception. So, relating specific brain measures using advanced neuroimaging to the change patients describe in their pain perception induced by peripheral or central sensitization (i.e. amplification), psychological or pharmacological mechanisms has tremendous value. Identifying where amplification or attenuation processes occur along the journey from injury to the brain (i.e. peripheral nerves, spinal cord, brainstem and brain) for an individual and relating these neural mechanisms to specific pain experiences, measures of pain relief, persistence of pain states, degree of injury and the subject's underlying genetics, has neuroscientific and potential diagnostic relevance. This is what neuroimaging has afforded – a better understanding and explanation of why someone’s pain is the way it is. We can go ‘behind the scenes’ of the subjective report to find out what key changes and mechanisms make up an individual’s particular pain experience. A key area of development has been pharmacological imaging where objective evidence of drugs reaching the target and working can be obtained. We even now understand the mechanisms of placebo analgesia – a powerful phenomenon known about for millennia. More recently, researchers have been investigating through brain imaging whether there is a pre-disposing vulnerability in brain networks towards developing chronic pain. So, advanced neuroimaging studies can powerfully aid explanation of a subject’s multidimensional pain experience, pain relief (analgesia) and even what makes them vulnerable to developing chronic pain. The application of this goes beyond the clinic and has relevance in courts of law, and other areas of society, such as in veterinary care. Relatively far less work has been directed at understanding what changes in the brain occur during altered states of consciousness induced either endogenously (e.g. sleep) or exogenously (e.g. anaesthesia). However, that situation is changing rapidly. Our recent multimodal neuroimaging work explores how anaesthetic agents produce altered states of consciousness such that perceptual experiences of pain and awareness are degraded. This is bringing us fascinating insights into the complex phenomenon of anaesthesia, consciousness and even the concept of self-hood. These topics will be discussed in my talk alongside my ‘side-story’ of life as a scientist combining academic leadership roles with doing science and raising a family.

Corticospinal neurons (CSN) are the cortical projection neurons that innervate the spinal cord and some brainstem targets with segmental precision to control voluntary movement of specific functional motor groups, limb sections, or individual digits, yet molecular regulation over CSN segmental target specificity is essentially unknown. CSN subpopulations exhibit striking axon targeting specificity from development into maturity: Evolutionarily newer rostrolateral CSN exclusively innervate bulbar-cervical targets (CSNBC-lat), while evolutionarily older caudomedial CSN (CSNmed) are more heterogeneous, with distinct subpopulations extending axons to either bulbar-cervical or thoraco-lumbar segments. The cervical cord, with its evolutionarily enhanced precision of forelimb movement, is innervated by multiple CSN subpopulations, suggesting inter-neuronal interactions in establishing corticospinal connectivity. I identify that Lumican, previously unrecognized in axon development, controls the specificity of cervical spinal cord innervation by CSN. Remarkably, Lumican, an extracellular matrix protein expressed by CSNBC-lat, non-cell-autonomously suppresses axon collateralization in the cervical cord by CSNmed. Intersectional viral labeling and mouse genetics further identify that Lumican controls axon collateralization by multiple subpopulations in caudomedial sensorimotor cortex. These results identify inter-axonal molecular crosstalk between CSN subpopulations as a novel mechanism controlling corticospinal connectivity and competitive specificity. Further, this mechanism has potential implications for evolutionary diversification of corticospinal circuitry with finer scale precision. "" Complementing this work, to comprehensively elucidate related axon projection mechanisms functioning at tips of growing CSN axons in vivo, I am currently applying experimental and analytic approaches recently developed in my postdoc lab (Poulopoulos*, Murphy*, Nature, 2019) to quantitatively and subcellularly “map” RNA and protein molecular machinery of subtype-specific growth cones, in parallel to their parent somata, isolated directly in vivo from developing subcerebral projection neurons (SCPN; the broader cortical output neuron population targeting both brainstem and spinal cord; includes CSN). I am investigating both normal development and GC-soma dysregulation with mutation of central CSN-SCPN transcriptional regulator Ctip2/Bcl11b.

Spontaneous firing, observed in many neurons, is often attributed to ion channel or network level noise. Cortical cells during slow wave sleep exhibit transitions between so called Up and Down states. In this sleep state, with limited sensory stimuli, neurons fire in the Up state. Spontaneous firing is also observed in slices of cholinergic interneurons, cerebellar Purkinje cells and even brainstem inspiratory neurons. In such in vitro preparations, where the functional relevance is long lost, neurons continue to display a rich repertoire of firing properties. It is perplexing that these neurons, instead of saving their energy during information downtime and functional irrelevance, are eager to fire. We propose that spontaneous firing is not a chance event but instead, a vital activity for the well-being of a neuron. We postulate that neurons, in anticipation of synaptic inputs, keep their ATP levels at maximum. As recovery from inputs requires most of the energy resources, neurons are ATP surplus and ADP scarce during synaptic quiescence. With ADP as the rate-limiting step, ATP production stalls in the mitochondria when ADP is low. This leads to toxic Reactive Oxygen Species (ROS) formation, which are known to disrupt many cellular processes. We hypothesize that spontaneous firing occurs at these conditions - as a release valve to spend energy and to restore ATP production, shielding the neuron against ROS. By linking a mitochondrial metabolism model to a conductance-based neuron model, we show that spontaneous firing depends on baseline ATP usage and on ATP-cost-per-spike. From our model, emerges a mitochondrial mediated homeostatic mechanism that provides a recipe for different firing patterns. Our findings, though mostly affecting intracellular dynamics, may have large knock-on effects on the nature of neural coding. Hitherto it has been thought that the neural code is optimised for energy minimisation, but this may be true only when neurons do not experience synaptic quiescence.

David Chalmers’s (1995) hard problem famously states: “It is widely agreed that experience arises from a physical basis, but we have no good explanation of why and how it so arises.” Thomas Nagel (1974) wrote something similar: “If we acknowledge that a physical theory of mind must account for the subjective character of experience, we must admit that no presently available conception gives us a clue about how this could be done.” This presentation will point the way towards the long-sought “good explanation” -- or at least it will provide “a clue”. I will make three points: (1) It is unfortunate that cognitive science took vision as its model example when looking for a ‘neural correlate of consciousness’ because cortical vision (like most cognitive processes) is not intrinsically conscious. There is not necessarily ‘something it is like’ to see. (2) Affective feeling, by contrast, is conscious by definition. You cannot feel something without feeling it. Moreover, affective feeling, generated in the upper brainstem, is the foundational form of consciousness: prerequisite for all the higher cognitive forms. (3) The functional mechanism of feeling explains why and how it cannot go on ‘in the dark’, free of any inner feel. Affect enables the organism to monitor deviations from its expected self-states in uncertain situations and thereby frees homeostasis from the limitations of automatism. As Nagel says, “An organism has conscious mental states if and only if there is something that it is like to be that organism—something it is like for the organism.” Affect literally constitutes the sentient subject.

Maintaining posture is a vital challenge for all freely-moving organisms. As animals grow, their relationship to destabilizing physical forces changes. How does the nervous system deal with this ongoing challenge? Vertebrates use highly conserved vestibular reflexes to stabilize the body. We established the larval zebrafish as a new model system to understand the development of the vestibular reflexes responsible for balance. In this talk, I will begin with the biophysical challenges facing baby fish as they learn to swim. I’ll briefly review published work by David Ehrlich, Ph.D., establishing a fundamental relationship between postural stability and locomotion. The bulk of the talk will highlight unpublished work by Kyla Hamling. She discovered that a small (~50) population of molecularly-defined brainstem neurons called vestibulo-spinal cells act as a nexus for postural development. Her loss-of-function experiments show that these neurons contribute more to postural stability as animals grow older. I’ll end with brief highlights from her ongoing work examining tilt-evoked responses of these neurons using 2-photon imaging and the consequences of downstream activity in the spinal cord using single-objective light-sheet (SCAPE) microscopy