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Neuroscience 2017

November 11 - November 15

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SfN’s 47th annual meeting, Neuroscience 2017, is the world’s largest neuroscience conference for scientists and physicians devoted to understanding the brain and nervous system.

SfN invites you to its renowned venue where neuroscientists collaborate and network with peers, learn from experts, explore the newest neuroscience tools and technologies, and discover great career opportunities.

Neuroscience 2017 is November 11-15 at the Walter E. Washington Convention Center. Join more than 30,000 colleagues from more than 80 countries at the world’s largest marketplace of ideas and tools for global neuroscience.

Visit us at booth #1934.

Cellular Dynamics Poster Presentations at Neuroscience 2017

Identification of Measurable Phenotypes Relevant to Alzheimer’s Disease Using Human iPSC-derived Neurons

Poster: 387.21
Monday, Nov 13 | 1:00 – 5:00 pm

Authors: Kile Mangan1, Lisa Harms1, Christian Kannemeier1, Kwi Hye Kim1, Benjamin M. Bader2, Konstantin Jügelt2, Olaf Schröder2, Beatriz Frietas1, Eugenia Jones1, and Coby Carlson1

  1. Cellular Dynamics International – a FUJIFILM company, Madison, WI
  2. NeuroProof GmbH – Rostock, Germany
Abstract

Alzheimer’s disease (AD) is a progressive neurodegenerative disease that results in gradual memory loss and impairment in the ability to learn or carry out daily tasks. The development of therapies for AD has been hindered by limited availability of relevant cell models for basic research and drug discovery. Using induced pluripotent stem cell (iPSC) technology, we have created an unlimited source of human neurons available for studying the mechanisms of AD progression and to streamline the identification of novel drug treatments for this disease. A hallmark of AD pathology is the development of plaques in the brain that contain toxic beta amyloid peptides (Abeta). Therefore, a key focus of AD research is to discern the specific contributions of Abeta to the disease.

We have taken two strategies to generate an iPSC-based “disease-in-a-dish” approach for modeling AD in vitro. The first is based on genome engineering of an apparently healthy normal iPSC line to introduce mutations in the gene coding for amyloid precursor protein (APP) and then create human neurons from genetically distinct samples. We rigorously tested the cell by high content imaging, PCR arrays, biomarker production, and multi-electrode array (MEA). Our data were in general agreement with results observed in other model systems for A673V (known to influence AD progression) and A673T (known to offer protection from the disease). Uniquely presented, however, functional assessment on MEA with multi-parametric analysis revealed the APP A673V mutant had a significantly different phenotype than A673T or the isogenic WT control.

Secondly, we have examined the effects of exogenous exposure to Abeta peptides. Addition of oligomeric Abeta(1-42) to GABAergic and glutamatergic neurons results in cytotoxicity as read out by ATP and LDH assays. Next, synchronous cultures of excitatory glutamatergic neurons – which can be analyzed on MEA to quantify bursting patterns, rates, intensities, and durations – display a dose-dependent decrease in network bursting prior to decay in firing rates and subsequent to cell death. Detailed evaluation of the burst structure and action potential morphology will be presented. Importantly, these alterations were not observed in control experiments with Abeta(1-40).

Our studies demonstrate the utility iPSC technology to create readily accessible human cell models for AD that recapitulate some of the functional neuronal phenotypes that are associated with this complex disease. Ultimately, the promise is that such gene-associated or in vitro disease models can be used to screen for compounds that rescue these phenotypes and significantly reduce the time and cost to develop new AD therapies and improve patient outcomes.


Human iPSC-derived Neurons Form Synchronously Bursting Cultures and Display a Seizurogenic Response to Excitatory Pharmacology

Poster: 381.28
Monday, Nov 13 | 1:00 – 5:00 pm

Authors: Kile Mangan, Lisa Harms, Elisabeth Enghofer, Christian Kannemeier, Coby Carlson, and Blake Anson
Cellular Dynamics International – a FUJIFILM company, Madison, WI

Abstract

The mammalian brain works appropriately only when there is a proper balance between excitation and inhibition. An imbalance in the ratio of excitatory-to- inhibitory neurons (referred to as the E/I ratio) is associated with numerous neurological abnormalities and deficits. Increased E/I ratios result in higher excitability and leads to prolonged neocortical circuit activity, stimulus hypersensitivity, cognitive impairments, and even epilepsy (Hagerman, et al. 2002; Gibson, et al., 2008; Zhang, et al. 2011). Of equal interest and importance, decreased E/I ratios result in a stronger inhibitory drive and have been linked to impaired social interactions, autistic behaviors, and mental retardation (Tabuchi, et al. 2007; Dani, et al. 2005). It is well-defined that during neuronal development that the E/I ratio changes and evolves over time, with excitation decreasing and inhibition increasing, and any deviations to this natural process may give rise to neurological disorders.

Despite the urgency to advance our understanding of the mechanisms that govern neural network formation and synchronous bursting behaviors, a major challenge in neuroscience research and the identification of new drugs is access to clinically-relevant cell models. The advent of induced pluripotent stem cell (iPSC) technology now grants us access to previously unattainable human neuronal cell types. Using this technology, we have generated iPSC-derived frontal cortical neurons of an appropriate, brain-similar E/I ratio (~80% excitation), that develop and display network-level, synapse-coupled, coordinated neuronal activity in vitro. Using multi-electrode arrays (MEA) to measure the electrophysiological activity of these glutamatergicneurons, we are able to capture periodical and synchronized bursting patterns and analyze these data via numerous parameters (including Poisson and ISI statistics). These neuronal cultures contain excitatory synapses (as shown by HOMER1 and synapsin co-staining) that modulate the synaptically-driven and synchronous bursting behaviors observed on the MEA. Importantly, these signals can be blocked by AP5 and DNQX.

In the interest of understanding how excitatory pharmacology alters network-level neuronal interactions, we investigated alterations in the regular bursting properties of glutamatergic neurons after addition of (0.003 to 300 micromolar dose response) various excitatory agents (e.g. bicuculline, chlorpromazine, pentylenetetrazol, amoxapine, 4-aminopyridine, and glutamic acid), common anti-epileptic drugs (e.g. ganaxalone and valproate), as well as a negative control (acetaminophen) and vehicle controls (e.g. ethanol and DMSO). Activity metrics displaying dose-dependent responses with pharmacology include: ‘single-channel’ Poisson bursts (rate, intensity and duration), ‘network-level’ ISI bursts (rate, intensity and duration), and synchrony measures. The presented data illustrates how human iPSC-technology can be leveraged to create an unprecedented investigatory space for understanding the intricacies of how excitatory synapses control network-connected populations of human neurons.


Calcium Handling Assays with Human iPSC-derived Neuronal Cell Types

Poster: 461.24
Tuesday, Nov 14 | 8:00 am – 12:00 pm

Authors: Kwi Hye Kim1 , Kile Mangan1 , Michael Hancock1 , Shouming Du2 , Kevin Oelstrom1 , and Coby Carlson1

  1. Cellular Dynamics International – a FUJIFILM company, Madison, WI
  2. Hamamatsu Corporation
Abstract

Human cell types differentiated from induced pluripotent stem cells (iPSC) offer an attractive source of cellular material for both toxicity screening and drug discovery because of the biologically relevant systems they can represent in vitro. In combination with cutting-edge instrumentation, iPSC- derived cells be used to explore functionality of human cells and to identify phenotypes for disease modeling. The FDSS uCell from Hamamatsu is a kinetic plate reader that is equipped with a high speed camera, an integrated dispenser head, and electrical field stimulation capability that enables intracellular ion measurement in live neuronal cell type. In this poster, we highlight the development of cell-based assays on the FDSS uCell to measure calcium handling in human iPSC-derived neurons. Specifically, we utilized GABAergic, glutamatergic, and dopaminergic neurons in both mono-culture and co-culture with iPSC-derived astrocytes. Addition of agonists that activate excitatory receptors, such as AMPA-R, Glutamate receptor, and NMDA-R, result in robust responses in calcium flux. Furthermore, we have also developed a method to measure spontaneous calcium oscillations, most importantly in iPSC- derived glutamatergic neurons, where these cells have been used to test the effects of various small-molecules as potentially seizurogenic. Finally, we have also leveraged the power of iPSC technology to illustrate phenotypic data from disease-specific or patient-derived samples, including a mutant KCNT1 channel that represents a rare but severe epilepsy syndrome. Taken together, these examples should help to create new avenues for safety assessment and toxicology studies, as well as serve as a template for future opportunities in modeling disease with human iPS cells.


Exploration of Mitochondrial Bioenergetics Using Human iPSC-derived Neurons

Poster: 671.2
Wednesday, Nov 15 | 8:00 am – 12:00 pm

Authors: Kwi Hye Kim,  Natsuyo Aoyama, Michael Hancock, Kile Mangan, and Coby Carlson
Cellular Dynamics International – a FUJIFILM company, Madison, WI

Abstract

The advent of induced pluripotent stem cell (iPSC) technology now grants the scientific community access to previously unattainable human cell types, specifically those from the brain. Moreover, iPSC technology enables us to compare cells from apparently healthy normal individuals to those cell types from disease-specific and patient-derived samples.

Neurological and neurodegenerative disorders are a global health concern. Many of these disease pathologies are thought to be driven by dysfunctional mitochondria creating an imbalance in cellular bioenergetics. The ability to measure discrete changes in cellular energy and metabolism in a real-time, label-free approach is possible with Seahorse XF technology.

We have used this instrument to assess the impact of numerous conditions (ranging from neuronal culture medium, time in culture, cell density, mitochondrial stress agents, and genetic mutations) on the bioenergetics profile human iPSC-derived cell types. Here we focus on development of the methods and protocols to evaluate mitochondrial stress in GABAergic, glutamatergic, dopaminergic, and motor neurons. Additionally, we have tested iPSC-derived astrocytes, macrophages, and skeletal muscle as each of these cell types are possibilities for co-culture to generate more complex and biologically relevant cell models.

Finally, establishing baseline assay signals for these various cell types derived from apparently healthy normal donors paves the way for disease modeling. Examples of neurons and astrocytes with the SOD1 G93A mutation will be presented.

iPSC-Derived Neurons Harboring a Known Epilepsy Mutation Display Known and Novel Electrophysiological Phenotypes

Authors: Mangan K1, Quaraishi I2, Zhang Y2, Carlson C1, McLachlan M1, Meline B1, Strouse A1, McMahon C1, Burke T1, Jones E1 and Kaczmarek L2

  1. Cellular Dynamics International – a FUJIFILM company, Madison, WI
  2. Yale University, School of Medicine, New Haven, CT
Abstract

Epilepsy is a disturbance in the electrical activity of the brain manifested via countless etiologies. 65 million individuals suffer from epilepsy and one-third of these individuals live with uncontrollable seizures because no known pharmacological treatment works for them. A portion of this population is accounted for by single-gene epilepsy disorders resulting from mutations within sodium, potassium or inhibitory channels. For example, the Slack gene (KCNT1) encodes a sodium-activated potassium channel that is very widely expressed in the brain. Mutations in this KCNT1 gene in humans presents with autosomal-dominant nocturnal frontal lobe epilepsy (ADNFLE), a disease marked by brief, but violent, seizures during sleep and devastating effects on intellectual function. Advances in personalized medicine is crucial for these types of diseases.

Central to this vision is induced pluripotent stem (iPS) cell technology, which provides a platform to expand our understanding of how single-gene mutations result in disease states. This approach illustrates and leverages the “disease-in-a-dish” iPSC-technology into phenotypic screening and drug development.

We have engineered and generated human cortical neurons harboring the KCNT1 {P924L}single-gene mutations, as well as the isogenic wild-type control match. This ability provides unprecedented access to in vitro models of alltypes of neurological disorders. Here we present functional data, via patch-clamp and multielectrode array (MEA) electrophysiological techniques, illustrating the known ‘gain-offunction’ ionotropic cellular-level fingerprint, which has previously been linked to this mutation, along with newly-discovered neuralnetwork level hyper-active phenotypes. We further show multiple examples that selective pharmacology can reverse these observed phenotypes. Collectively, our results illustrate how human iPS cells can be model disease states and be leveraged in the personal medicine space.

Details

Start:
November 11
End:
November 15
Website:
https://www.sfn.org/annual-meeting/neuroscience-2017

Organizer

Society for Neuroscience

Venue

Walter E. Washington Convention Center
801 Mt Vernon Pl NW
Washington, DC 20001 United States
Website:
http://www.dcconvention.com/Venues/ConventionCenter.aspx