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Zlatic Lab / Publications
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18 Publications

Showing 1-10 of 18 results
08/09/17 | The complete connectome of a learning and memory centre in an insect brain.
Eichler K, Li F, Litwin-Kumar A, Park Y, Andrade I, Schneider-Mizell CM, Saumweber T, Huser A, Eschbach C, Gerber B, Fetter RD, Truman JW, Priebe CE, Abbott LF, Thum AS, Zlatic M, Cardona A
Nature. 2017 Aug 09;548(7666):175-182. doi: 10.1038/nature23455

Associating stimuli with positive or negative reinforcement is essential for survival, but a complete wiring diagram of a higher-order circuit supporting associative memory has not been previously available. Here we reconstruct one such circuit at synaptic resolution, the Drosophila larval mushroom body. We find that most Kenyon cells integrate random combinations of inputs but that a subset receives stereotyped inputs from single projection neurons. This organization maximizes performance of a model output neuron on a stimulus discrimination task. We also report a novel canonical circuit in each mushroom body compartment with previously unidentified connections: reciprocal Kenyon cell to modulatory neuron connections, modulatory neuron to output neuron connections, and a surprisingly high number of recurrent connections between Kenyon cells. Stereotyped connections found between output neurons could enhance the selection of learned behaviours. The complete circuit map of the mushroom body should guide future functional studies of this learning and memory centre.

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07/01/17 | The Ol1mpiad: concordance of behavioural faculties of stage 1 and stage 3 Drosophila larvae.
Almeida-Carvalho MJ, Berh D, Braun A, Chen Y, Eichler K, Eschbach C, Fritsch PM, Gerber B, Hoyer N, Jiang X, Kleber J, Klämbt C, König C, Louis M, Michels B, Miroschnikow A, Mirth C, Miura D, Niewalda T, Otto N, Paisios E, Pankratz MJ, Petersen M, Ramsperger N, Randel N, Risse B, Saumweber T, Schlegel P, Schleyer M, Soba P, Sprecher SG, Tanimura T, Thum AS, Toshima N, Truman JW, Yarali A, Zlatic M
The Journal of Experimental Biology. 2017 Jul 01;220(Pt 13):2452-2475. doi: 10.1242/jeb.156646

Mapping brain function to brain structure is a fundamental task for neuroscience. For such an endeavour, the Drosophila larva is simple enough to be tractable, yet complex enough to be interesting. It features about 10,000 neurons and is capable of various taxes, kineses and Pavlovian conditioning. All its neurons are currently being mapped into a light-microscopical atlas, and Gal4 strains are being generated to experimentally access neurons one at a time. In addition, an electron microscopic reconstruction of its nervous system seems within reach. Notably, this electron microscope-based connectome is being drafted for a stage 1 larva - because stage 1 larvae are much smaller than stage 3 larvae. However, most behaviour analyses have been performed for stage 3 larvae because their larger size makes them easier to handle and observe. It is therefore warranted to either redo the electron microscopic reconstruction for a stage 3 larva or to survey the behavioural faculties of stage 1 larvae. We provide the latter. In a community-based approach we called the Ol1mpiad, we probed stage 1 Drosophila larvae for free locomotion, feeding, responsiveness to substrate vibration, gentle and nociceptive touch, burrowing, olfactory preference and thermotaxis, light avoidance, gustatory choice of various tastants plus odour-taste associative learning, as well as light/dark-electric shock associative learning. Quantitatively, stage 1 larvae show lower scores in most tasks, arguably because of their smaller size and lower speed. Qualitatively, however, stage 1 larvae perform strikingly similar to stage 3 larvae in almost all cases. These results bolster confidence in mapping brain structure and behaviour across developmental stages.

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05/09/17 | Semiparametric spectral modeling of the Drosophila connectome.
Priebe CE, Park Y, Tang M, Athreya A, Lyzinski V, Vogelstein JT, Qin Y, Cocanougher B, Eichler K, Zlatic M, Cardona A
arXiv. 2017 May 9:1705.03297

We present semiparametric spectral modeling of the complete larval Drosophila mushroom body connectome. Motivated by a thorough exploratory data analysis of the network via Gaussian mixture modeling (GMM) in the adjacency spectral embedding (ASE) representation space, we introduce the latent structure model (LSM) for network modeling and inference. LSM is a generalization of the stochastic block model (SBM) and a special case of the random dot product graph (RDPG) latent position model, and is amenable to semiparametric GMM in the ASE representation space. The resulting connectome code derived via semiparametric GMM composed with ASE captures latent connectome structure and elucidates biologically relevant neuronal properties.

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04/19/17 | Pavlovian conditioning of larval Drosophila: an illustrated, multilingual, hands-on manual for odor-taste associative learning in maggots.
Michels B, Saumweber T, Biernacki R, Thur J, Glasgow RD, Schleyer M, Chen Y, Eschbach C, Stocker RF, Toshima N, Tanimura T, Louis M, Arias-Gil G, Marescotti M, Benfenati F, Gerber B
Frontiers in Behavioral Neuroscience. 2017 Apr 19;11:45. doi: 10.3389/fnbeh.2017.00045

Larval Drosophila offer a study case for behavioral neurogenetics that is simple enough to be experimentally tractable, yet complex enough to be worth the effort. We provide a detailed, hands-on manual for Pavlovian odor-reward learning in these animals. Given the versatility of Drosophila for genetic analyses, combined with the evolutionarily shared genetic heritage with humans, the paradigm has utility not only in behavioral neurogenetics and experimental psychology, but for translational biomedicine as well. Together with the upcoming total synaptic connectome of the Drosophila nervous system and the possibilities of single-cell-specific transgene expression, it offers enticing opportunities for research. Indeed, the paradigm has already been adopted by a number of labs and is robust enough to be used for teaching in classroom settings. This has given rise to a demand for a detailed, hands-on manual directed at newcomers and/or at laboratory novices, and this is what we here provide. The paradigm and the present manual have a unique set of features: • The paradigm is cheap, easy, and robust; • The manual is detailed enough for newcomers or laboratory novices; • It briefly covers the essential scientific context; • It includes sheets for scoring, data analysis, and display; • It is multilingual: in addition to an English version we provide German, French, Japanese, Spanish and Italian language versions as well. The present manual can thus foster science education at an earlier age and enable research by a broader community than has been the case to date.

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03/31/17 | Facilitating Neuron-Specific Genetic Manipulations in Drosophila Using a Split GAL4 Repressor.
Dolan M, Luan H, Shropshire WC, Sutcliffe B, Cocanougher B, Scott RL, Frechter S, Zlatic M, Jefferis GS, White BH
Genetics. 2017 Mar 31;206(2):775-84. doi: 10.1534/genetics.116.199687

Efforts to map neural circuits have been galvanized by the development of genetic technologies that permit the manipulation of targeted sets of neurons in the brains of freely behaving animals. The success of these efforts relies on the experimenter's ability to target arbitrarily small subsets of neurons for manipulation, but such specificity of targeting cannot routinely be achieved using existing methods. In Drosophila melanogaster, a widely used technique for refined cell-type specific manipulation is the Split GAL4 system, which augments the targeting specificity of the binary GAL4-UAS system by making GAL4 transcriptional activity contingent upon two enhancers, rather than one. To permit more refined targeting, we introduce here the "Killer Zipper" (KZip(+)), a suppressor that makes Split GAL4 targeting contingent upon a third enhancer. KZip(+) acts by disrupting both the formation and activity of Split GAL4 heterodimers, and we show how this added layer of control can be used to selectively remove unwanted cells from a Split GAL4 expression pattern or to subtract neurons of interest from a pattern to determine their requirement in generating a given phenotype. To facilitate application of the KZip(+) technology, we have developed a versatile set of LexAop-KZip(+) fly lines that can be used directly with the large number of LexA driver lines with known expression patterns. The Killer Zipper significantly sharpens the precision of neuronal genetic control available in Drosophila and may be extended to other organisms where Split GAL4-like systems are used.

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10/05/16 | Competitive disinhibition mediates behavioral choice and sequences in Drosophila.
Jovanic T, Schneider-Mizell CM, Shao M, Masson J, Denisov G, Fetter RD, Mensh BD, Truman JW, Cardona A, Zlatic M
Cell. 2016 Oct 5;167(3):858-70. doi: 10.1016/j.cell.2016.09.009

Even a simple sensory stimulus can elicit distinct innate behaviors and sequences. During sensorimotor decisions, competitive interactions among neurons that promote distinct behaviors must ensure the selection and maintenance of one behavior, while suppressing others. The circuit implementation of these competitive interactions is still an open question. By combining comprehensive electron microscopy reconstruction of inhibitory interneuron networks, modeling, electrophysiology, and behavioral studies, we determined the circuit mechanisms that contribute to the Drosophila larval sensorimotor decision to startle, explore, or perform a sequence of the two in response to a mechanosensory stimulus. Together, these studies reveal that, early in sensory processing, (1) reciprocally connected feedforward inhibitory interneurons implement behavioral choice, (2) local feedback disinhibition provides positive feedback that consolidates and maintains the chosen behavior, and (3) lateral disinhibition promotes sequence transitions. The combination of these interconnected circuit motifs can implement both behavior selection and the serial organization of behaviors into a sequence.

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02/10/16 | Four Individually Identified Paired Dopamine Neurons Signal Reward in Larval Drosophila.
Rohwedder A, Wenz NL, Stehle B, Huser A, Yamagata N, Zlatic M, Truman JW, Tanimoto H, Saumweber T, Gerber B, Thum AS
Current Biology : CB. 2016 Feb 10:. doi: 10.1016/j.cub.2016.01.012

Dopaminergic neurons serve multiple functions, including reinforcement processing during associative learning [1-12]. It is thus warranted to understand which dopaminergic neurons mediate which function. We study larval Drosophila, in which only approximately 120 of a total of 10,000 neurons are dopaminergic, as judged by the expression of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine biosynthesis [5, 13]. Dopaminergic neurons mediating reinforcement in insect olfactory learning target the mushroom bodies, a higher-order "cortical" brain region [1-5, 11, 12, 14, 15]. We discover four previously undescribed paired neurons, the primary protocerebral anterior medial (pPAM) neurons. These neurons are TH positive and subdivide the medial lobe of the mushroom body into four distinct subunits. These pPAM neurons are acutely necessary for odor-sugar reward learning and require intact TH function in this process. However, they are dispensable for aversive learning and innate behavior toward the odors and sugars employed. Optogenetical activation of pPAM neurons is sufficient as a reward. Thus, the pPAM neurons convey a likely dopaminergic reward signal. In contrast, DL1 cluster neurons convey a corresponding punishment signal [5], suggesting a cellular division of labor to convey dopaminergic reward and punishment signals. On the level of individually identified neurons, this uncovers an organizational principle shared with adult Drosophila and mammals [1-4, 7, 9, 10] (but see [6]). The numerical simplicity and connectomic tractability of the larval nervous system [16-19] now offers a prospect for studying circuit principles of dopamine function at unprecedented resolution.

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09/03/15 | Identification of Inhibitory Premotor Interneurons Activated at a Late Phase in a Motor Cycle during Drosophila Larval Locomotion.
Itakura Y, Kohsaka H, Ohyama T, Zlatic M, Pulver SR, Nose A
PLoS One. 2015 Sep 03;10(9):e0136660. doi: 10.1371/journal.pone.0136660

Rhythmic motor patterns underlying many types of locomotion are thought to be produced by central pattern generators (CPGs). Our knowledge of how CPG networks generate motor patterns in complex nervous systems remains incomplete, despite decades of work in a variety of model organisms. Substrate borne locomotion in Drosophila larvae is driven by waves of muscular contraction that propagate through multiple body segments. We use the motor circuitry underlying crawling in larval Drosophila as a model to try to understand how segmentally coordinated rhythmic motor patterns are generated. Whereas muscles, motoneurons and sensory neurons have been well investigated in this system, far less is known about the identities and function of interneurons. Our recent study identified a class of glutamatergic premotor interneurons, PMSIs (period-positive median segmental interneurons), that regulate the speed of locomotion. Here, we report on the identification of a distinct class of glutamatergic premotor interneurons called Glutamatergic Ventro-Lateral Interneurons (GVLIs). We used calcium imaging to search for interneurons that show rhythmic activity and identified GVLIs as interneurons showing wave-like activity during peristalsis. Paired GVLIs were present in each abdominal segment A1-A7 and locally extended an axon towards a dorsal neuropile region, where they formed GRASP-positive putative synaptic contacts with motoneurons. The interneurons expressed vesicular glutamate transporter (vGluT) and thus likely secrete glutamate, a neurotransmitter known to inhibit motoneurons. These anatomical results suggest that GVLIs are premotor interneurons that locally inhibit motoneurons in the same segment. Consistent with this, optogenetic activation of GVLIs with the red-shifted channelrhodopsin, CsChrimson ceased ongoing peristalsis in crawling larvae. Simultaneous calcium imaging of the activity of GVLIs and motoneurons showed that GVLIs' wave-like activity lagged behind that of motoneurons by several segments. Thus, GVLIs are activated when the front of a forward motor wave reaches the second or third anterior segment. We propose that GVLIs are part of the feedback inhibition system that terminates motor activity once the front of the motor wave proceeds to anterior segments.

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04/20/15 | A multilevel multimodal circuit enhances action selection in Drosophila.
Ohyama T, Schneider-Mizell CM, Fetter RD, Aleman JV, Franconville R, Rivera-Alba M, Mensh BD, Branson KM, Simpson JH, Truman JW, Cardona A, Zlatic M
Nature. 2015 Apr 20;520(7549):633-9. doi: 10.1038/nature14297

Natural events present multiple types of sensory cues, each detected by a specialized sensory modality. Combining information from several modalities is essential for the selection of appropriate actions. Key to understanding multimodal computations is determining the structural patterns of multimodal convergence and how these patterns contribute to behaviour. Modalities could converge early, late or at multiple levels in the sensory processing hierarchy. Here we show that combining mechanosensory and nociceptive cues synergistically enhances the selection of the fastest mode of escape locomotion in Drosophila larvae. In an electron microscopy volume that spans the entire insect nervous system, we reconstructed the multisensory circuit supporting the synergy, spanning multiple levels of the sensory processing hierarchy. The wiring diagram revealed a complex multilevel multimodal convergence architecture. Using behavioural and physiological studies, we identified functionally connected circuit nodes that trigger the fastest locomotor mode, and others that facilitate it, and we provide evidence that multiple levels of multimodal integration contribute to escape mode selection. We propose that the multilevel multimodal convergence architecture may be a general feature of multisensory circuits enabling complex input–output functions and selective tuning to ecologically relevant combinations of cues.

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02/13/15 | Labeling of active neural circuits in vivo with designed calcium integrators.
Fosque BF, Sun Y, Dana H, Yang C, Ohyama T, Tadross MR, Patel R, Zlatic M, Kim DS, Ahrens MB, Jayaraman V, Looger LL, Schreiter ER
Science. 2015 Feb 13;347(6223):755-60. doi: 10.1126/science.1260922

The identification of active neurons and circuits in vivo is a fundamental challenge in understanding the neural basis of behavior. Genetically encoded calcium (Ca(2+)) indicators (GECIs) enable quantitative monitoring of cellular-resolution activity during behavior. However, such indicators require online monitoring within a limited field of view. Alternatively, post hoc staining of immediate early genes (IEGs) indicates highly active cells within the entire brain, albeit with poor temporal resolution. We designed a fluorescent sensor, CaMPARI, that combines the genetic targetability and quantitative link to neural activity of GECIs with the permanent, large-scale labeling of IEGs, allowing a temporally precise "activity snapshot" of a large tissue volume. CaMPARI undergoes efficient and irreversible green-to-red conversion only when elevated intracellular Ca(2+) and experimenter-controlled illumination coincide. We demonstrate the utility of CaMPARI in freely moving larvae of zebrafish and flies, and in head-fixed mice and adult flies.

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