Publications by authors named "Amanda Crocker"

14 Publications

  • Page 1 of 1

Molecular Systems Biology of Neurodevelopmental Disorders, Rett Syndrome as an Archetype.

Front Integr Neurosci 2019 17;13:30. Epub 2019 Jul 17.

Rare Neurological Diseases (Private Research Institution), Atlanta, GA, United States.

Neurodevelopmental disorders represent a challenging biological and medical problem due to their genetic and phenotypic complexity. In many cases, we lack the comprehensive understanding of disease mechanisms necessary for targeted therapeutic development. One key component that could improve both mechanistic understanding and clinical trial design is reliable molecular biomarkers. Presently, no objective biological markers exist to evaluate most neurodevelopmental disorders. Here, we discuss how systems biology and "omic" approaches can address the mechanistic and biomarker limitations in these afflictions. We present heuristic principles for testing the potential of systems biology to identify mechanisms and biomarkers of disease in the example of Rett syndrome, a neurodevelopmental disorder caused by a well-defined monogenic defect in methyl-CpG-binding protein 2 (MECP2). We propose that such an approach can not only aid in monitoring clinical disease severity but also provide a measure of target engagement in clinical trials. By deepening our understanding of the "big picture" of systems biology, this approach could even help generate hypotheses for drug development programs, hopefully resulting in new treatments for these devastating conditions.
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http://dx.doi.org/10.3389/fnint.2019.00030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6650571PMC
July 2019

Studying alcohol use disorder using Drosophila melanogaster in the era of 'Big Data'.

Behav Brain Funct 2019 Apr 16;15(1). Epub 2019 Apr 16.

Program in Neuroscience, Middlebury College, Middlebury, VT, 05753, USA.

Our understanding of the networks of genes and protein functions involved in Alcohol Use Disorder (AUD) remains incomplete, as do the mechanisms by which these networks lead to AUD phenotypes. The fruit fly (Drosophila melanogaster) is an efficient model for functional and mechanistic characterization of the genes involved in alcohol behavior. The fly offers many advantages as a model organism for investigating the molecular and cellular mechanisms of alcohol-related behaviors, and for understanding the underlying neural circuitry driving behaviors, such as locomotor stimulation, sedation, tolerance, and appetitive (reward) learning and memory. Fly researchers are able to use an extensive variety of tools for functional characterization of gene products. To understand how the fly can guide our understanding of AUD in the era of Big Data we will explore these tools, and review some of the gene networks identified in the fly through their use, including chromatin-remodeling, glial, cellular stress, and innate immunity genes. These networks hold great potential as translational drug targets, making it prudent to conduct further research into how these gene mechanisms are involved in alcohol behavior.
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http://dx.doi.org/10.1186/s12993-019-0159-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6469124PMC
April 2019

Systems Analysis of the 22q11.2 Microdeletion Syndrome Converges on a Mitochondrial Interactome Necessary for Synapse Function and Behavior.

J Neurosci 2019 05 4;39(18):3561-3581. Epub 2019 Mar 4.

Departments of Cell Biology,

Neurodevelopmental disorders offer insight into synaptic mechanisms. To unbiasedly uncover these mechanisms, we studied the 22q11.2 syndrome, a recurrent copy number variant, which is the highest schizophrenia genetic risk factor. We quantified the proteomes of 22q11.2 mutant human fibroblasts from both sexes and mouse brains carrying a 22q11.2-like defect, Molecular ontologies defined mitochondrial compartments and pathways as some of top ranked categories. In particular, we identified perturbations in the SLC25A1-SLC25A4 mitochondrial transporter interactome as associated with the 22q11.2 genetic defect. Expression of SLC25A1-SLC25A4 interactome components was affected in neuronal cells from schizophrenia patients. Furthermore, hemideficiency of the SLC25A1 or SLC25A4 orthologues, dSLC25A1-sea and dSLC25A4-sesB, affected synapse morphology, neurotransmission, plasticity, and sleep patterns. Our findings indicate that synapses are sensitive to partial loss of function of mitochondrial solute transporters. We propose that mitoproteomes regulate synapse development and function in normal and pathological conditions in a cell-specific manner. We address the central question of how to comprehensively define molecular mechanisms of the most prevalent and penetrant microdeletion associated with neurodevelopmental disorders, the 22q11.2 microdeletion syndrome. This complex mutation reduces gene dosage of ∼63 genes in humans. We describe a disruption of the mitoproteome in 22q11.2 patients and brains of a 22q11.2 mouse model. In particular, we identify a network of inner mitochondrial membrane transporters as a hub required for synapse function. Our findings suggest that mitochondrial composition and function modulate the risk of neurodevelopmental disorders, such as schizophrenia.
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http://dx.doi.org/10.1523/JNEUROSCI.1983-18.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6495129PMC
May 2019

Cell-Type-Specific Transcriptome Analysis in the Drosophila Mushroom Body Reveals Memory-Related Changes in Gene Expression.

Cell Rep 2016 05 5;15(7):1580-1596. Epub 2016 May 5.

Princeton Neuroscience Institute, Princeton University, Princeton, NJ 08544, USA; Department of Molecular Biology, Princeton University, Princeton, NJ 08544, USA. Electronic address:

Learning and memory formation in Drosophila rely on a network of neurons in the mushroom bodies (MBs). Whereas numerous studies have delineated roles for individual cell types within this network in aspects of learning or memory, whether or not these cells can also be distinguished by the genes they express remains unresolved. In addition, the changes in gene expression that accompany long-term memory formation within the MBs have not yet been studied by neuron type. Here, we address both issues by performing RNA sequencing on single cell types (harvested via patch pipets) within the MB. We discover that the expression of genes that encode cell surface receptors is sufficient to identify cell types and that a subset of these genes, required for sensory transduction in peripheral sensory neurons, is not only expressed within individual neurons of the MB in the central brain, but is also critical for memory formation.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5047377PMC
http://dx.doi.org/10.1016/j.celrep.2016.04.046DOI Listing
May 2016

Interaction between sleep and metabolism in Drosophila with altered octopamine signaling.

J Biol Chem 2012 Sep 24;287(39):32406-14. Epub 2012 Jul 24.

Department of Neuroscience, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Sleep length and metabolic dysfunction are correlated, but the causal relationship between these processes is unclear. Octopamine promotes wakefulness in the fly by acting through the insulin-producing cells (IPCs) in the fly brain. To determine if insulin signaling mediates the effects of octopamine on sleep:wake behavior, we assayed flies in which insulin signaling activity was genetically altered. We found that increasing insulin signaling does not promote wake, nor does insulin appear to mediate the wake-promoting effects of octopamine. Octopamine also affects metabolism in invertebrate species, including, as we show here, Drosophila melanogaster. Triglycerides are decreased in mutants with compromised octopamine signaling and elevated in flies with increased activity of octopaminergic neurons. Interestingly, this effect is mediated at least partially by insulin, suggesting that effects of octopamine on metabolism are independent of its effects on sleep. We further investigated the relative contribution of metabolic and sleep phenotypes to the starvation response of flies with altered octopamine signaling. Hyperactivity (indicative of foraging) induced by starvation was elevated in octopamine receptor mutants, despite their high propensity for sleep, indicating that their metabolic state dictates their behavioral response under these conditions. Moreover, flies with increased octopamine signaling do not suppress sleep in response to starvation, even though they are normally hyper-aroused, most likely because of their high triglyceride levels. Together, these data suggest that observed correlations between sleep and metabolic phenotypes can result from shared molecular pathways rather than causality, and environmental conditions can lead to the dominance of one phenotype over the other.
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http://dx.doi.org/10.1074/jbc.M112.360875DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3463357PMC
September 2012

The central clock neurons regulate lipid storage in Drosophila.

PLoS One 2011 18;6(5):e19921. Epub 2011 May 18.

Department of Neuroscience, The University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

A proper balance of lipid breakdown and synthesis is essential for achieving energy homeostasis as alterations in either of these processes can lead to pathological states such as obesity. The regulation of lipid metabolism is quite complex with multiple signals integrated to control overall triglyceride levels in metabolic tissues. Based upon studies demonstrating effects of the circadian clock on metabolism, we sought to determine if the central clock cells in the Drosophila brain contribute to lipid levels in the fat body, the main nutrient storage organ of the fly. Here, we show that altering the function of the Drosophila central clock neurons leads to an increase in fat body triglycerides. We also show that although triglyceride levels are not affected by age, they are increased by expression of the amyloid-beta protein in central clock neurons. The effect on lipid storage seems to be independent of circadian clock output as changes in triglycerides are not always observed in genetic manipulations that result in altered locomotor rhythms. These data demonstrate that the activity of the central clock neurons is necessary for proper lipid storage.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0019921PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3097222PMC
November 2011

Genetic analysis of sleep.

Genes Dev 2010 Jun;24(12):1220-35

Howard Hughes Medical institute, Department of Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, USA.

Almost 20 years ago, the gene underlying fatal familial insomnia was discovered, and first suggested the concept that a single gene can regulate sleep. In the two decades since, there have been many advances in the field of behavioral genetics, but it is only in the past 10 years that the genetic analysis of sleep has emerged as an important discipline. Major findings include the discovery of a single gene underlying the sleep disorder narcolepsy, and identification of loci that make quantitative contributions to sleep characteristics. The sleep field has also expanded its focus from mammalian model organisms to Drosophila, zebrafish, and worms, which is allowing the application of novel genetic approaches. Researchers have undertaken large-scale screens to identify new genes that regulate sleep, and are also probing questions of sleep circuitry and sleep function on a molecular level. As genetic tools continue to be refined in each model organism, the genes that support a specific function in sleep will become more apparent. Thus, while our understanding of sleep still remains rudimentary, rapid progress is expected from these recently initiated studies.
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http://dx.doi.org/10.1101/gad.1913110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2885658PMC
June 2010

Identification of a neural circuit that underlies the effects of octopamine on sleep:wake behavior.

Neuron 2010 Mar;65(5):670-81

Department of Neuroscience, University of Pennsylvania Medical School, Philadelphia, PA 19104, USA.

An understanding of sleep requires the identification of distinct cellular circuits that mediate the action of specific sleep:wake-regulating molecules, but such analysis has been very limited. We identify here a circuit that underlies the wake-promoting effects of octopamine in Drosophila. Using MARCM, we identified the ASM cells in the medial protocerebrum as the wake-promoting octopaminergic cells. We then blocked octopamine signaling in random areas of the fly brain and mapped the postsynaptic effect to insulin-secreting neurons of the pars intercerebralis (PI). These PI neurons show altered potassium channel function as well as an increase in cAMP in response to octopamine, and genetic manipulation of their electrical excitability alters sleep:wake behavior. Effects of octopamine on sleep:wake are mediated by the cAMP-dependent isoform of the OAMB receptor. These studies define the cellular and molecular basis of octopamine action and suggest that the PI is a sleep:wake-regulating neuroendocrine structure like the mammalian hypothalamus.
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http://dx.doi.org/10.1016/j.neuron.2010.01.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2862355PMC
March 2010

The effects of caffeine on sleep in Drosophila require PKA activity, but not the adenosine receptor.

J Neurosci 2009 Sep;29(35):11029-37

Division of Sleep Medicine, Department of Neurology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Caffeine is one of the most widely consumed stimulants in the world and has been proposed to promote wakefulness by antagonizing function of the adenosine A2A receptor. Here, we show that chronic administration of caffeine reduces and fragments sleep in Drosophila and also lengthens circadian period. To identify the mechanisms underlying these effects of caffeine, we first generated mutants of the only known adenosine receptor in flies (dAdoR), which by sequence is most similar to the mammalian A2A receptor. Mutants lacking dAdoR have normal amounts of baseline sleep, as well as normal homeostatic responses to sleep deprivation. Surprisingly, these mutants respond normally to caffeine. On the other hand, the effects of caffeine on sleep and circadian rhythms are mimicked by a potent phosphodiesterase inhibitor, IBMX (3-isobutyl-1-methylxanthine). Using in vivo fluorescence resonance energy transfer imaging, we find that caffeine induces widespread increase in cAMP levels throughout the brain. Finally, the effects of caffeine on sleep are blocked in flies that have reduced neuronal PKA activity. We suggest that chronic administration of caffeine promotes wakefulness in Drosophila, at least in part, by inhibiting cAMP phosphodiesterase activity.
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http://dx.doi.org/10.1523/JNEUROSCI.1653-09.2009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2757164PMC
September 2009

Octopamine regulates sleep in drosophila through protein kinase A-dependent mechanisms.

J Neurosci 2008 Sep;28(38):9377-85

Department of Neuroscience, Howard Hughes Medical Institute, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

Sleep is a fundamental process, but its regulation and function are still not well understood. The Drosophila model for sleep provides a powerful system to address the genetic and molecular mechanisms underlying sleep and wakefulness. Here we show that a Drosophila biogenic amine, octopamine, is a potent wake-promoting signal. Mutations in the octopamine biosynthesis pathway produced a phenotype of increased sleep, which was restored to wild-type levels by pharmacological treatment with octopamine. Moreover, electrical silencing of octopamine-producing cells decreased wakefulness, whereas excitation of these neurons promoted wakefulness. Because protein kinase A (PKA) is a putative target of octopamine signaling and is also implicated in Drosophila sleep, we investigated its role in the effects of octopamine on sleep. We found that decreased PKA activity in neurons rendered flies insensitive to the wake-promoting effects of octopamine. However, this effect of PKA was not exerted in the mushroom bodies, a site previously associated with PKA action on sleep. These studies identify a novel pathway that regulates sleep in Drosophila.
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http://dx.doi.org/10.1523/JNEUROSCI.3072-08a.2008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2742176PMC
September 2008

Protecting motor neurons from toxic insult by antagonism of adenosine A2a and Trk receptors.

J Neurosci 2006 Sep;26(36):9250-63

Department of Neurology, Children's Hospital of Philadelphia, Joseph Stokes Jr. Research Institute, Philadelphia, Pennsylvania 19104, USA.

The death of motor neurons in amyotrophic lateral sclerosis (ALS) is thought to result from the interaction of a variety of factors including excitotoxicity, accumulation of toxic proteins, and abnormal axonal transport. Previously, we found that the susceptibility of motor neurons to excitotoxic insults can be limited by inhibiting signals evoked by brain-derived neurotrophic factor (BDNF) activation of the receptor tyrosine kinase B (TrkB). Here we show that this can be achieved by direct kinase inhibition or by blockade of a transactivation pathway that uses adenosine A2a receptors and src-family kinases (SFKs). Downstream signaling cascades (such as mitogen-activated protein kinase and phosphatidylinositol-3 kinase) are inhibited by these blockers. In addition to protecting motor neurons from excitotoxic insult, these agents also prevent toxicity that follows from the expression of mutant proteins (G85R superoxide dismutase 1; G59S p150(glued)) that cause familial motor neuron disease. TrkB, adenosine A2a receptors, and SFKs associate into complexes in lipid raft and nonlipid raft membranes and the signaling from lipids rafts may be particularly important because their disruption by cholesterol depletion blocks the ability of BDNF to render motor neurons vulnerable to insult. The neuroprotective versatility of Trk antagonism suggests that it may have broad utility in the treatment of ALS patients.
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http://dx.doi.org/10.1523/JNEUROSCI.1856-06.2006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6674510PMC
September 2006

Sleep in Drosophila is regulated by adult mushroom bodies.

Nature 2006 Jun;441(7094):757-60

Center for Sleep and Respiratory Neurobiology, University of Pennsylvania Medical School, Philadelphia, Pennsylvania 19104, USA.

Sleep is one of the few major whole-organ phenomena for which no function and no underlying mechanism have been conclusively demonstrated. Sleep could result from global changes in the brain during wakefulness or it could be regulated by specific loci that recruit the rest of the brain into the electrical and metabolic states characteristic of sleep. Here we address this issue by exploiting the genetic tractability of the fruitfly, Drosophila melanogaster, which exhibits the hallmarks of vertebrate sleep. We show that large changes in sleep are achieved by spatial and temporal enhancement of cyclic-AMP-dependent protein kinase (PKA) activity specifically in the adult mushroom bodies of Drosophila. Other manipulations of the mushroom bodies, such as electrical silencing, increasing excitation or ablation, also alter sleep. These results link sleep regulation to an anatomical locus known to be involved in learning and memory.
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http://dx.doi.org/10.1038/nature04811DOI Listing
June 2006

Afferents to the orexin neurons of the rat brain.

J Comp Neurol 2006 Feb;494(5):845-61

Department of Neurology, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA.

Emotions, stress, hunger, and circadian rhythms all promote wakefulness and behavioral arousal. Little is known about the pathways mediating these influences, but the orexin-producing neurons of the hypothalamus may play an essential role. These cells heavily innervate many wake-promoting brain regions, and mice lacking the orexin neurons have narcolepsy and fail to rouse in response to hunger (Yamanaka et al. [2003] Neuron 38:701-713). To identify the afferents to the orexin neurons, we first injected a retrograde tracer into the orexin neuron field of rats. Retrogradely labeled neurons were abundant in the allocortex, claustrum, lateral septum, bed nucleus of the stria terminalis, and in many hypothalamic regions including the preoptic area, dorsomedial nucleus, lateral hypothalamus, and posterior hypothalamus. Retrograde labeling in the brainstem was generally more modest, but labeling was strong in the periaqueductal gray matter, dorsal raphe nucleus, and lateral parabrachial nucleus. Injection of an anterograde tracer confirmed that most of these regions directly innervate the orexin neurons, with some of the heaviest input coming from the lateral septum, preoptic area, and posterior hypothalamus. In addition, hypothalamic regions preferentially innervate orexin neurons in the medial and perifornical parts of the field, but most projections from the brainstem target the lateral part of the field. Inputs from the suprachiasmatic nucleus are mainly relayed via the subparaventricular zone and dorsomedial nucleus. These observations suggest that the orexin neurons may integrate a variety of interoceptive and homeostatic signals to increase behavioral arousal in response to hunger, stress, circadian signals, and autonomic challenges.
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http://dx.doi.org/10.1002/cne.20859DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2259441PMC
February 2006

Behavioral state instability in orexin knock-out mice.

J Neurosci 2004 Jul;24(28):6291-300

Department of Neurology, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02115, USA.

Narcolepsy is caused by a lack of orexin (hypocretin), but the physiologic process that underlies the sleepiness of narcolepsy is unknown. Using orexin knock-out (KO) mice as a model of narcolepsy, we critically tested the three leading hypotheses: poor circadian control of sleep and wakefulness, inadequate activation of arousal regions, or abnormal sleep homeostasis. Compared with wild-type (WT) littermates, orexin KO mice had essentially normal amounts of sleep and wake, but wake and non-rapid eye movement (NREM) bouts were very brief, with many more transitions between all behavioral states. In constant darkness, orexin KO mice had normal amplitude and timing of sleep-wake rhythms, providing no evidence for disordered circadian control. When placed in a new, clean cage, both groups of mice remained awake for approximately 45 min, demonstrating that, even in the absence of orexin, fundamental arousal regions can be engaged to produce sustained wakefulness. After depriving mice of sleep for 2-8 hr, orexin KO mice recovered their NREM and rapid eye movement sleep deficits at comparable rates and to the same extent as WT mice, with similar increases in EEG delta power, suggesting that their homeostatic control of sleep is normal. These experiments demonstrate that the fragmented wakefulness of orexin deficiency is not a consequence of abnormal sleep homeostasis, poor circadian control, or defective fundamental arousal systems. Instead, the fragmented behavior of orexin KO mice may be best described as behavioral state instability, with apparently low thresholds to transition between states.
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http://dx.doi.org/10.1523/JNEUROSCI.0586-04.2004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6729542PMC
July 2004