A new chemogenetic method that enables functional neural circuit analysis

Neurons within the brain are intricately organized and interconnected to form neural circuits which work collectively and specifically to control behaviour and physiology. Understanding how these neuronal circuits function in the mammalian brain and how they are dysregulated in disease is a major goal in neuroscience research. Two techniques, chemo- and opto-genetics, have revolutionized neuroscience as they have allowed precise exploration of the involvement of specific neurons in modulating particular behaviours and physiology by exogenous drug or light activation, respectively. However, both opto- and chemo-genetics lack the ability to define the inputs to the neuron which drives the behavior and this circuit information is essential for understanding behavior and its dysregulation in disease.

In collaborative studies with Professor Andrew Allen in the Department of Physiology and Dr Stuart McDougall at the Florey we have developed a new chemogenetic tool to understand neural circuits. Our research, which was recently published in Cell Reports (1), utilizes an adaptation of a chemogenetic system using the insect peptide allatostatin-3 (Ast-3). Both Ast-3 and its specific allatostatin receptor (AstR) are inert in the mammalian brain. Expression of AstR in specific neurons, and then delivery of exogenous Ast-3 peptide has been used in mammals to study neuronal function. Our development was to exploit the fact that the Ast-3 peptide can be genetically encoded. We developed a recombinant viral vector that enables expression of bioactive Ast-3, its trafficking to the synapse and release in response to neural activity to inhibit neurons expressing the AstR. We have therefore developed an enhanced, highly specific chemogenetic approach whereby genetic expression of the activating ligand enables definition of the inputs to neurons expressing the AstR and hence understanding of the circuits controlling the behaviours associated with the specific neurons.

Current studies, available for student projects, are focused on expanding this chemogenetic methodology to make the Ast-3 construct cell-specific, based upon neurochemical phenotype or its axonal projection and Ast-3 expression inducible using a tetracycline transactivator approach. Additionally, we will develop a novel excitatory version of this novel system. Projects will involve molecular cloning of the viral constructs and extensive validation of their functionality in cell systems in the Bathgate lab; detailed synaptic physiology to examine transgenic peptide release with whole cell neural recording in the McDougall lab; and in vivo validation both anatomically and physiologically using well-characterized neural systems in Prof Andrew Allen’s laboratory. Honours, Masters and PhD projects are available.

Relevant Publications:

1. Ngo H, Melo MR, Layfield S, Connelly AA, Bassi JK, Xie L, Menuet C, McDougall SJ, Bathgate RAD*, Allen AM* (2020) A novel chemogenetic tool for functional neural circuits analysis. Cell reports 32, 108139

2. Ma S, Allocca G, Ong-Pålsson EKE, Hawkes D, McDougall SJ, Williams SJ, Bathgate RAD, Gundlach AL. (2016) Nucleus incertus promotes cortical desynchronization and behavioral arousal. Brain Structure and Function 222: 515-537.

3. Ganella DE, Callander GE, Ma S, Gundlach AL, Bathgate RAD (2013) Modulation of feeding by chronic rAAV expression of a relaxin-3 peptide agonist in rat hypothalamus. Gene Therapy 20, 703-71

4. Callander GE, Ganella DE, Ma S, Wimmer V, Gundlach AL, Thomas WG, Bathgate RAD (2012) Silencing relaxin-3 in nucleus incertus of adult rodents: a viral vector-based approach to investigate neuropeptide function. PLoS One. 7, e42300.

Research team

Collaborators

Professor Andrew Allen (Department of Physiology)