Model Systems Core Services

Sets of human IDD cellular models are derived for each project by reprogramming patient-derived biomaterials to generate induced PSC lines, and by genome engineering of both WT and IDD subject-derived PSCs. The CTC works with investigators to recruit subjects and obtain deidentified biomaterials, which are provided to the CMU for this purpose. Reprogramming and genome engineering is either performed by the CMU or through collaboration with the Genome Engineering and induced Pluripotency Center @WUSTL. The CMU also derives molecular tools to manipulate the activity of IDD genes, which are used in combination with these PSC models.

PSC models are differentiated into a range of IDD-relevant neuronal and organoid types and used to perform cellular and molecular phenotyping, including assessment of:

• cellular physiology, including altered proliferative and apoptotic indices, ER or oxidative stress, and/or cellular metabolism.
• neural progenitor specification, maintenance, and progenitor cell type identity.
• neuronal differentiation, organoid patterning, neuronal cell type identity, and maturation.
• neuronal morphology, neurite extension, synaptogenesis, and migration.
• neuronal function, assessed using electrophysiology and/or multi-electrode array, in pure neuronal populations, organoids, or co-cultures, is performed in collaboration with Dr. Huettner in the NPhyS.
• Genome-wide transcriptomic and/or epigenomic analyses are used to define molecular mechanisms, pathways, and processes altered in IDD models, including acute effects and direct targets. RNA or chromatin sample preparation is performed by the CMU, with sample sequencing performed through the Genome Technology Access Center @WUSTL.
• Targeted or high-throughput screening and other applications.

The workflow above defines cell autonomous, intermediate phenotypes linked to patient affectation.  These cells are useful for small molecule and chemical screening to identify suppressors, and to identify leads for developing interventions for IDD. The CMU performs some of this screening in collaboration with the High-Throughput Screening Core @WUSTL. These models can also be used for many other applications, such as high throughput annotation of variant pathogenicity by complementation assay, to compare phenotypes resulting from many variants of uncertain pathogenicity in an IDD gene.

The directors of the CMUs at WUSTL and University of Wisconsin, Drs. Kroll and Anita Bhattacharyya, co-lead the Cross-IDDRC Cellular Models Group. This group engages CMUs at all 14 US IDDRCs in collaborative projects, some of which focus on cross-IDDRC calibration, benchmarking, and standardization of methods for IDD cellular phenotyping, to ensure reproducibility and enable data meta-analysis across the network. The Cellular Models Group is also building both a biorepository and a data repository for IDD cellular models under study at all 14 IDDRCs. These repositories will enable access to IDD models generated by CMUs throughout the network, and will link these models with accompanying genomic, transcriptomic, and epigenomic data. Banking of these IDD models and data will utilize cross-IDDRC calibrated methods, making these repositories valuable for data meta-analyses and model resource sharing by all 14 IDDRCs. Such meta-analyses can be informative in defining convergent mechanisms that contribute to IDDs involving many different genetic contributors to patient risk.

The ABS offers over 50 individual mouse behavioral tests within the following categories:

1. Approach/Avoidance (Anxiety-Like)
2. Circadian Rhythm
3. Developmental Trajectories
4. Feeding Behavior
5. Learning and Memory (spatial and non-spatial)
6. Repetitive, Restricted Behaviors
7. Reward Behavior
8. Sensory/Motor Functions
9. Social Behaviors

For rats, we offer behavioral tests within the following categories:

1. Approach/Avoidance (Anxiety-Like)
2. Learning and Memory (spatial and non-spatial)
3. Sensory/Motor Function
4. Social Behaviors

Our team of animal behavior experts can provide consultation on experimental issues such as sex effects, background strain, littermate controls, non-behavioral influences on performance, and statistical power. Consultation will include recommending certain tests for specific models and cost estimates for the work involved, as well as referrals to other relevant cores.

The ABS has acquired a wide range of instrumentation to better characterize phenotypes and treatment effects including:

1. DigiGait and CatWalk gait analysis systems
2. Mouse and Rat Morris Water Maze Pools
3. Force-plate Actometers
4. Accoustic Startle/Prepulse Inhibition Chambers and Program
5. Fear Conditioning Chambers and Program
6. Behavior Sound- and Scent-Attenuating Chambers for various tests

And more!

Training on our equipment and protocols can be provided as a service if requested.

The ABS is currently developing and integrating machine learning into our behavioral tests and analyses with the goal of allowing us to offer streamlined access to high-throughput machine learning tools for IDD researchers.  Machine learning techniques will allow for the automated acquisition and precise processing of subtle behaviors during testing through the use of computer vision applications, which will expand our phenotyping capabilities.

We are excitedly building the necessary infrastructure required for high-throughput computer vision and machine learning behavior quantification, including:

1) Building new custom behavioral data acquisition systems designed specifically for the advanced video acquisition required for computer vision platforms.

2) Acquiring high-powered machines necessary to build and process computer vision and machine learning models.

3) Generating computer vision and machine learning models for automated quantification of gait and motor behaviors, stereotypies and repetitive patterns of behavior, and social behaviors.

Basic histopathology approaches for light microscopic evaluation include perfusion fixation, embedding, sectioning and staining of CNS tissues.  The NPS also provides investigator training in these methods and in research design, data analysis and interpretation. Additional procedures offered include DeOlmos cupric silver staining, in situ hybridization, and western blotting.

The NPS has experience with a wide variety of immunohistochemical methods including: activated caspase-3, pERK, TUNEL, NeuN, GFAP, MBP, Ki-67, PCNA, Nestin, BrdU, NR1, GAD, calbindin, ChAT.  The NPS  will develop additional approaches as need arises. Previous work included the development of new methods to assess the developmental neuropathology produced by Zika virus infection, anesthetic drugs, and glucocorticoids. 

The NPS also conducts and/or assists investigators in quantitative tissue analyses using unbiased stereology and unbiased sampling of the brain or brain regions of interest. The NPS is equipped with digital imaging microscopes with electronically driven stages and Microbrightfield software specifically designed to facilitate unbiased stereological quantitative analysis. The NPS also provides ultrastructural analysis of pathological changes in the developing brain. Both Drs. Noguchi and Farber have many years of experience in using the electron microscope to investigate a variety of neuropathological phenomena including viral infection, excitoxicity, and apoptosis.

EEG: Numerous studies have documented the co-existence of seizures with disorders of IDD. In order to evaluate these and other paroxysmal behaviors, the NPhyS established and offers a rodent epilepsy monitoring unit with the capacity to monitor 36 mice using continuous video-EEG monitoring.  Using this technique, the NPhyS has characterized seizures in multiple models of IDD, such as in mouse models of tuberous sclerosis complex, neuronal ceroid lipofuscinosis, and other genetic IDDs.  The NPhyS also developed and offers technically innovative methods to allow serial video-EEG monitoring longitudinally in preweaning neonatal mice. In addition, NPhys performs both qualitative and quantitative (e.g., spike counts, spectral analysis) assessments of abnormalities in interictal background EEG activity, which provide a neurophysiological correlate of encephalopathy;

Seizure threshold testing: This is performed using convulsant drugs, such as pentylenetetrazol or kainate in rodents that do not have spontaneous seizures;

Polysomnograms: Sleep disorders are common in IDD and can exacerbate its symptoms. Using a combination of EEG and EMG, the NPhyS performs sleep studies that characterize the sleep-wake cycle, including the percentage of times in awake, non-REM, and REM states and power spectra analysis. The NPhyS has recently used these methods to identify sleep abnormalities in a mouse model of neurofibromatosis, tuberous sclerosis, and aging.

Somatosensory evoked potentials: Sensory abnormalities are also characteristic of some neurodevelopmental disorders. The NPhyS offers newly developed methods for stimulating median nerve in rodent forepaw and stably recording evoked potentials over somatosensory cortex with epidural electrodes;

Slice and Cellular Electrophysiology: Extracellular field potential recordings and single cell/intracellular recordings are powerful methods for assessing synaptic function and plasticity in neuronal networks and investigating the cellular basis and mechanisms of learning deficits and other brain dysfunction in IDDs.  The NPhyS offers consultation, training, and equipment use to investigators in several fully-equipped electrophysiology rigs available in the MSC for extracellular field potential recording in brain slices and whole-cell patch-clamp recording in slices and cultured cells.

Human PSCs:  Inasmuch as neurons derived from human PSCs are difficult to work with and characterize, Dr. Huettner, a noted expert in this field, works with Dr. Kroll of the Cellular Models Unit to assess IDD patient-derived pure neuronal populations, organoids, or co-cultures using electrophysiology and/or multi-electrode array.