Hoffman Lab Research
What are the biological mechanisms underlying
How can we develop novel, mechanism-based
In the Hoffman lab, our goal is to understand the basic biological mechanisms underlying neurodevelopmental disorders, in particular autism spectrum disorders (ASD). To do this, we study the function of genes that are strongly associated with autism in humans to determine how disruption of these genes alters brain development and the neural circuits underlying simple behaviors. Our long-term goal is to use this gene-based approach to identify relevant biological pathways and novel pharmacological treatments that target these pathways.
The zebrafish system
We use zebrafish as a model to understand the function of genes involved in autism. There are three main advantages of studying zebrafish:
First, zebrafish have external fertilization and development and transparent embryos, which allows us to observe key processes in nervous system development in vivo.
Brain images of 4-day-old zebrafish larvae showing wild-type fish and mutants of the autism risk gene, CNTNAP2, which encodes a cell adhesion molecule. Images show inhibitory (green) and excitatory neurons (magenta) and axon tracts (cyan) (Hoffman et al. 2016, Neuron).
Second, zebrafish have large progenies, which allows us to conduct high-throughput behavioral assays and pharmacological screens. We can study the locomotor activity of hundreds of zebrafish larvae simultaneously using an automated tracking system.
The locomotor activity of zebrafish larvae can be tracked and quantified using an automated assay.
Third, the introduction of CRISPR/Cas9 technology has advanced our ability to target many genes of interest. Our laboratory is using CRISPRs to generate multiple lines of zebrafish mutants in genes that are strongly associated with autism risk.
In the Hoffman lab, we aim to understand how disruption of ASD risk genes alters specific neural pathways contributing to autism. Whole-exome sequencing in large-scale human studies has led to a rapid advance in the ability to identify with confidence genes that are likely to contribute to autism risk.
We study structural and behavioral phenotypes in zebrafish mutants of autism risk genes to identify common biological pathways involving these genes.
Autism Risk Genes and Brain Development
It has been proposed that imbalance in excitatory and inhibitory neurotransmission is a mechanism underlying ASD. To study the simultaneous development of these cell populations, we use transgenic lines (gifts from the labs of M. Ekker and J. Fetcho) that co-label these populations and examine their development in the brains of zebrafish mutants of autism risk genes.
Using this approach, we found that zebrafish mutants of the gene, CNTNAP2, which is associated with a syndrome of autism and epilepsy in individuals from the Old Order Amish population, have selective deficits in GABAergic neurons, particularly in the forebrain at 4 days post fertilization (Hoffman et al., 2016, Neuron). In contrast, we did not find specific regional deficits in glutamatergic neurons or abnormalities in the structure of the axon scaffold at this stage. We are interested in analyzing the extent to which other ASD risk genes affect excitatory and inhibitory neurons.
Zebrafish cntnap2 mutants have deficits in GABAergic neurons (green) particularly in the forebrain (Hoffman et al. 2016, Neuron).
High-throughput Pharmaco-Behavioral Analysis
We use an automated system to track the locomotor activity of zebrafish larvae. This system provides a quantitative readout of the behavior of the fish. We use this system to determine how disruption of ASD risk genes in zebrafish mutants affects the neural circuits underlying simple locomotor behaviors. We found that zebrafish mutants of cntnap2 display a specific behavioral phenotype of nighttime hyperactivity (Hoffman et al., 2016, Neuron). By comparing the activity profile of cntnap2 mutants to the profiles of wild-type fish exposed to over 500 psychoactive drugs, we predicted drugs that could exacerbate or suppress the mutant behavioral phenotype. We found that cntnap2 mutants show increased sensitivity to psychoactive agents that antagonize GABA and NMDA receptors. We found that drugs with estrogenic activity were able to reverse nighttime hyperactivity in cntnap2 mutants (Hoffman et al., 2016, Neuron). Working in collaboration with Jason Rihel’s lab at UCL, we are continuing this work to investigate common pharmaco-behavioral pathways that are disrupted in zebrafish mutants of other ASD risk genes.
The cntnap2 mutant behavioral profile clusters with psychoactive compounds that increase nighttime activity in wild-type fish (Hoffman et al. 2016, Neuron).
The Hoffman lab works with collaborators at Yale University and researchers at other universities to advance our understanding of the biology of autism. Some of our collaborators are:
Thomas Fernandez, Yale Child Study Center
Antonio Giraldez, Yale Department of Genetics
Abha Gupta, Yale Department of Pediatrics
Jason Rihel, University College London
Matthew State, UCSF
Steve Wilson, University College London