Neuroendocrine systems in animals maintain organismal homeostasis and regulate stress response. Although a great deal of work has been done on the neuropeptides and hormones that are released and act ...on target organs in the periphery, the synaptic inputs onto these neuroendocrine outputs in the brain are less well understood. Here, we use the transmission electron microscopy reconstruction of a whole central nervous system in the
larva to elucidate the sensory pathways and the interneurons that provide synaptic input to the neurosecretory cells projecting to the endocrine organs. Predicted by network modeling, we also identify a new carbon dioxide-responsive network that acts on a specific set of neurosecretory cells and that includes those expressing corazonin (Crz) and diuretic hormone 44 (Dh44) neuropeptides. Our analysis reveals a neuronal network architecture for combinatorial action based on sensory and interneuronal pathways that converge onto distinct combinations of neuroendocrine outputs.
In pursuit of food, hungry animals mobilize significant energy resources and overcome exhaustion and fear. How need and motivation control the decision to continue or change behavior is not ...understood. Using a single fly treadmill, we show that hungry flies persistently track a food odor and increase their effort over repeated trials in the absence of reward suggesting that need dominates negative experience. We further show that odor tracking is regulated by two mushroom body output neurons (MBONs) connecting the MB to the lateral horn. These MBONs, together with dopaminergic neurons and Dop1R2 signaling, control behavioral persistence. Conversely, an octopaminergic neuron, VPM4, which directly innervates one of the MBONs, acts as a brake on odor tracking by connecting feeding and olfaction. Together, our data suggest a function for the MB in internal state-dependent expression of behavior that can be suppressed by external inputs conveying a competing behavioral drive.
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•Hunger motivates persistent food odor tracking even without reward•Two synaptically connected MBONs, -γ1pedc>αβ and -α2sc, regulate odor tracking•Octopamine neurons connect feeding and counteract MBON and odor tracking•Dopaminergic neurons and Dop1R2 signaling promote persistent tracking
What drives behavioral persistence versus quitting? Sayin et al. propose that circuit modules in the fly’s learning center and dopamine drive gradually increasing food odor tracking, which can be efficiently suppressed by extrinsic, but directly innervating, feeding-related neuromodulatory neurons.
Steroid hormones play key roles in development, growth, and reproduction in various animal phyla 1. The insect steroid hormone, ecdysteroid, coordinates growth and maturation, represented by molting ...and metamorphosis 2. In Drosophila melanogaster, the prothoracicotropic hormone (PTTH)-producing neurons stimulate peak levels of ecdysteroid biosynthesis for maturation 3. Additionally, recent studies on PTTH signaling indicated that basal levels of ecdysteroid negatively affect systemic growth prior to maturation 4–8. However, it remains unclear how PTTH signaling is regulated for basal ecdysteroid biosynthesis. Here, we report that Corazonin (Crz)-producing neurons regulate basal ecdysteroid biosynthesis by affecting PTTH neurons. Crz belongs to gonadotropin-releasing hormone (GnRH) superfamily, implying an analogous role in growth and maturation 9. Inhibition of Crz neuronal activity increased pupal size, whereas it hardly affected pupariation timing. This phenotype resulted from enhanced growth rate and a delay in ecdysteroid elevation during the mid-third instar larval (L3) stage. Interestingly, Crz receptor (CrzR) expression in PTTH neurons was higher during the mid- than the late-L3 stage. Silencing of CrzR in PTTH neurons increased pupal size, phenocopying the inhibition of Crz neuronal activity. When Crz neurons were optogenetically activated, a strong calcium response was observed in PTTH neurons during the mid-L3, but not the late-L3, stage. Furthermore, we found that octopamine neurons contact Crz neurons in the subesophageal zone (SEZ), transmitting signals for systemic growth. Together, our results suggest that the Crz-PTTH neuronal axis modulates ecdysteroid biosynthesis in response to octopamine, uncovering a regulatory neuroendocrine system in the developmental transition from growth to maturation.
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•Crz neurons contact both PG cells and PTTH neurons•The Crz-PTTH neuronal axis controls growth via basal ecdysteroid biosynthesis•PTTH neurons respond to Crz neurons during the mid-L3 stage with CrzR expression•Octopamine neurons contact with Crz neurons in the SEZ, regulating systemic growth
The insect steroid hormone, ecdysteroid, coordinates growth and maturation. Imura et al. identify that the Crz-PTTH neuronal axis controls growth by regulating basal ecdysteroid production in response to octopamine neurons. Crz/CrzR is homologous to GnRH/GnRHR, which is required for puberty, implying an analogous role in growth and maturation.
Animals exhibit innate and learned preferences for temperature and humidity—conditions critical for their survival and reproduction. Leveraging a whole-brain electron microscopy volume, we studied ...the adult Drosophila melanogaster circuitry associated with antennal thermo- and hygrosensory neurons. We have identified two new target glomeruli in the antennal lobe, in addition to the five known ones, and the ventroposterior projection neurons (VP PNs) that relay thermo- and hygrosensory information to higher brain centers, including the mushroom body and lateral horn, seats of learned and innate behavior. We present the first connectome of a thermo- and hygrosensory neuropil, the lateral accessory calyx (lACA), by reconstructing neurons downstream of heating- and cooling-responsive VP PNs. A few mushroom body-intrinsic neurons solely receive thermosensory input from the lACA, while most receive additional olfactory and thermo- and/or hygrosensory PN inputs. Furthermore, several classes of lACA-associated neurons form a local network with outputs to other brain neuropils, suggesting that the lACA serves as a hub for thermo- and hygrosensory circuitry. For example, DN1a neurons link thermosensory PNs in the lACA to the circadian clock via the accessory medulla. Finally, we survey strongly connected downstream partners of VP PNs across the protocerebrum; these include a descending neuron targeted by dry-responsive VP PNs, meaning that just two synapses might separate hygrosensory inputs from motor circuits. These data provide a comprehensive first- and second-order layer analysis of Drosophila thermo- and hygrosensory systems and an initial survey of third-order neurons that could directly modulate behavior.
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•Two novel thermo- and/or hygrosensory glomeruli in the fly antennal lobe•First complete set of thermosensory and hygrosensory projection neurons•First connectome for a thermo- and hygrosensory neuropil•Third-order thermo- and hygrosensory neurons, including link to circadian clock
Marin et al. use connectomics and genetics for comprehensive identification of temperature and humidity sensory neurons in the Drosophila brain. They reconstruct all projections to higher brain areas and select higher-order targets, including the mushroom body lateral accessory calyx, linking thermosensation to memory and the circadian clock.
Different types of Drosophila dopaminergic neurons (DANs) reinforce memories of unique valence and provide state-dependent motivational control 1. Prior studies suggest that the compartment ...architecture of the mushroom body (MB) is the relevant resolution for distinct DAN functions 2, 3. Here we used a recent electron microscope volume of the fly brain 4 to reconstruct the fine anatomy of individual DANs within three MB compartments. We find the 20 DANs of the γ5 compartment, at least some of which provide reward teaching signals, can be clustered into 5 anatomical subtypes that innervate different regions within γ5. Reconstructing 821 upstream neurons reveals input selectivity, supporting the functional relevance of DAN sub-classification. Only one PAM-γ5 DAN subtype γ5(fb) receives direct recurrent feedback from γ5β′2a mushroom body output neurons (MBONs) and behavioral experiments distinguish a role for these DANs in memory revaluation from those reinforcing sugar memory. Other DAN subtypes receive major, and potentially reinforcing, inputs from putative gustatory interneurons or lateral horn neurons, which can also relay indirect feedback from MBONs. We similarly reconstructed the single aversively reinforcing PPL1-γ1pedc DAN. The γ1pedc DAN inputs mostly differ from those of γ5 DANs and they cluster onto distinct dendritic branches, presumably separating its established roles in aversive reinforcement and appetitive motivation 5, 6. Tracing also identified neurons that provide broad input to γ5, β′2a, and γ1pedc DANs, suggesting that distributed DAN populations can be coordinately regulated. These connectomic and behavioral analyses therefore reveal further complexity of dopaminergic reinforcement circuits between and within MB compartments.
•Nanoscale anatomy reveals additional subtypes of rewarding dopaminergic neurons•Connectomics reveals input specificity to subtypes of dopaminergic neurons•Axon morphology implies dopaminergic neurons provide subcompartment-level function•Unique dopaminergic subtypes serve aversive memory extinction and sugar learning
Otto et al. use electron microscope resolution connectomics to describe the structure and organization of neurons providing synaptic input to functionally discrete subtypes of dopaminergic neurons. The nanoscale anatomy reveals further anatomical and functional specialization of dopaminergic neurons, which is confirmed with behavioral experiments.
The clinical use of the chemotherapeutic doxorubicin (Dox) is limited by cardiotoxic side-effects. One of the early Dox effects is induction of a sarcoplasmic reticulum (SR) Ca2+ leak. The chaperone ...Glucose regulated protein 78 (GRP78) is important for Ca2+ homeostasis in the endoplasmic reticulum (ER)-the organelle corresponding to the SR in non-cardiomyocytes-and has been shown to convey resistance to Dox in certain tumors. Our aim was to investigate the effect of cardiac GRP78 gene transfer on Ca2+ dependent signaling, cell death, cardiac function and survival in clinically relevant in vitro and in vivo models for Dox cardiotoxicity.By using neonatal cardiomyocytes we could demonstrate that Dox induced Ca2+ dependent Ca2+ /calmodulin-dependent protein kinase II (CaMKII) activation is one of the factors involved in Dox cardiotoxicity by promoting apoptosis. Furthermore, we found that adeno-associated virus (AAV) mediated GRP78 overexpression partly protects neonatal cardiomyocytes from Dox induced cell death by modulating Ca2+ dependent pathways like the activation of CaMKII, phospholamban (PLN) and p53 accumulation. Most importantly, cardiac GRP78 gene therapy in mice treated with Dox revealed improved diastolic function (dP/dtmin) and survival after Dox treatment. In conclusion, our results demonstrate for the first time that Ca2+ dependent CaMKII activation fosters Dox cardiomyopathy and provide additional insight into possible mechanisms by which GRP78 overexpression protects cardiomyocytes from Doxorubicin toxicity.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The behavioral response to a sensory stimulus may depend on both learned and innate neuronal representations. How these circuits interact to produce appropriate behavior is unknown. In Drosophila, ...the lateral horn (LH) and mushroom body (MB) are thought to mediate innate and learned olfactory behavior, respectively, although LH function has not been tested directly. Here we identify two LH cell types (PD2a1 and PD2b1) that receive input from an MB output neuron required for recall of aversive olfactory memories. These neurons are required for aversive memory retrieval and modulated by training. Connectomics data demonstrate that PD2a1 and PD2b1 neurons also receive direct input from food odor-encoding neurons. Consistent with this, PD2a1 and PD2b1 are also necessary for unlearned attraction to some odors, indicating that these neurons have a dual behavioral role. This provides a circuit mechanism by which learned and innate olfactory information can interact in identified neurons to produce appropriate behavior.
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•Specific Drosophila lateral horn neurons mediate innate attraction to food odors•The same neurons receive plastic odor information from the mushroom body•Recall after associative learning depends on reduced drive to lateral horn neurons•Connectomics circuit for integration of learned and innate odor representations
Sensory stimuli can engage both learned and innate behaviors. Dolan et al. identify neurons in Drosophila that directly integrate unlearned and plastic odor representations; they are required for innate approach to food odors but also learned aversive recall.
Central mechanisms by which specific motor programs are selected to achieve meaningful behaviors are not well understood. Using electrophysiological recordings from pharyngeal nerves upon central ...activation of neurotransmitter-expressing cells, we show that distinct neuronal ensembles can regulate different feeding motor programs. In behavioral and electrophysiological experiments, activation of 20 neurons in the brain expressing the neuropeptide hugin, a homolog of mammalian neuromedin U, simultaneously suppressed the motor program for food intake while inducing the motor program for locomotion. Decreasing hugin neuropeptide levels in the neurons by RNAi prevented this action. Reducing the level of hugin neuronal activity alone did not have any effect on feeding or locomotion motor programs. Furthermore, use of promoter-specific constructs that labeled subsets of hugin neurons demonstrated that initiation of locomotion can be separated from modulation of its motor pattern. These results provide insights into a neural mechanism of how opposing motor programs can be selected in order to coordinate feeding and locomotive behaviors.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Motor systems can be functionally organized into effector organs (muscles and glands), the motor neurons, central pattern generators (CPG) and higher control centers of the brain. Using genetic and ...electrophysiological methods, we have begun to deconstruct the motor system driving Drosophila larval feeding behavior into its component parts. In this paper, we identify distinct clusters of motor neurons that execute head tilting, mouth hook movements, and pharyngeal pumping during larval feeding. This basic anatomical scaffold enabled the use of calcium-imaging to monitor the neural activity of motor neurons within the central nervous system (CNS) that drive food intake. Simultaneous nerve- and muscle-recordings demonstrate that the motor neurons innervate the cibarial dilator musculature (CDM) ipsi- and contra-laterally. By classical lesion experiments we localize a set of CPGs generating the neuronal pattern underlying feeding movements to the subesophageal zone (SEZ). Lesioning of higher brain centers decelerated all feeding-related motor patterns, whereas lesioning of ventral nerve cord (VNC) only affected the motor rhythm underlying pharyngeal pumping. These findings provide a basis for progressing upstream of the motor neurons to identify higher regulatory components of the feeding motor system.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The increase in protein activity and upregulation of G-protein coupled receptor kinase 2 (GRK2) is a hallmark of cardiac stress and heart failure. Inhibition of GRK2 improved cardiac function and ...survival and diminished cardiac remodeling in various animal heart failure models. The aim of the present study was to investigate the effects of GRK2 on cardiac hypertrophy and dissect potential molecular mechanisms. In mice we observed increased GRK2 mRNA and protein levels following transverse aortic constriction (TAC). Conditional GRK2 knockout mice showed attenuated hypertrophic response with preserved ventricular geometry 6 weeks after TAC operation compared to wild-type animals. In isolated neonatal rat ventricular cardiac myocytes stimulation with angiotensin II and phenylephrine enhanced GRK2 expression leading to enhanced signaling via protein kinase B (PKB or Akt), consecutively inhibiting glycogen synthase kinase 3 beta (GSK3β), such promoting nuclear accumulation and activation of nuclear factor of activated T-cells (NFAT). Cardiac myocyte hypertrophy induced by in vitro GRK2 overexpression increased the cytosolic interaction of GRK2 and phosphoinositide 3-kinase γ (PI3Kγ). Moreover, inhibition of PI3Kγ as well as GRK2 knock down prevented Akt activation resulting in halted NFAT activity and reduced cardiac myocyte hypertrophy. Our data show that enhanced GRK2 expression triggers cardiac hypertrophy by GRK2-PI3Kγ mediated Akt phosphorylation and subsequent inactivation of GSK3β, resulting in enhanced NFAT activity.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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