Summary: The researchers developed ConVERGD, a tool to precisely manipulate specific cell subpopulations, improving studies of cell diversity. The study demonstrates the utility of ConVERGD by identifying a norepinephrine neuron subpopulation associated with anxiety. This innovative approach could impact research and treatment in a variety of fields.
Key facts:
- A precision instrument: ConVERGD enables precise targeting and manipulation of cell subpopulations.
- Application in neurology: Used to identify a norepinephrine neuronal subpopulation implicated in anxiety.
- Wide impact: Potential applications beyond neuroscience, benefiting various research areas.
source: St. Jude Children’s Research Hospital
As gene sequencing technologies become more powerful, our understanding of cellular diversity grows in parallel. This prompted scientists from the Children’s Research Hospital St. Jude to create a tool to improve the ease and precision with which researchers can study specific subpopulations of cells.
The tool, called Conditional Viral Expression by Ribozyme Guided Degradation (ConVERGD), allows researchers to specifically access these subsets of cells and precisely manipulate them based on multiple cell characteristics.
ConVERGD offers numerous advantages over existing cross-sectional expression platforms by accommodating more complex genetic payloads and increased adaptability.
The researchers demonstrated the utility of ConVERGD by examining a previously unidentified subpopulation of norepinephrine neurons. The work demonstrates the significant impact that studies of cell subpopulations can have on basic research and healthcare.
The findings were published today in Nature Neurology.
Same cell type, different functions
About Lindsay Schwartz, PhD, St. Jude Department of Developmental Neurobiology, necessity led to invention as she explored the landscape of neuronal cells and specifically the neuronal cells that produce norepinephrine.
“Norepinephrine neurons are thought to be just one type of neuron. But when activated in the brain, they can cause many different types of behavior, such as enhancing attention and memory formation or triggering a stress response or a fight-or-flight response,” Schwartz said.
“But if only one kind of neuron releases that one molecule, then how does it make you do different things?”
To investigate such questions requires the ability to selectively interrogate cell subpopulations with extreme biases. To that end, Schwartz found that all attempts using current practices were failing.
“We didn’t go into this project thinking we were going to create a new tool, but it seemed like a need in the community.”
Improving current cell subpopulation targeting technology
Targeting subpopulations of cells requires passing them through several genetic filters. These cross-sectional filters interrogate what genes cells express and what pathways and connections they make, analyzing different subpopulations so that researchers can focus on a select group of isolated cells.
The use of adeno-associated virus (AAV)-based reporter tools that can deliver genetic material into specific cells with high fidelity is an ideal approach to implement these intersection filters.
These reporter tools are used to label or monitor gene expression and protein localization in specific cells or regions. However, they can be complex to design and offer limited space within them.
“One of our main goals was to design a tool where the gene of interest is only expressed when it’s passed with multiple functions, but it’s really easy for end users to modify and put in whatever genes they want,” Schwartz explained.
Robust ribozymes offer next-generation specificity
Schwartz and first author Alex Hughes, PhD, a graduate of the St. Jude Graduate School of Biomedical Sciences, now at the Allen Institute for Brain Sciences, used two separate technologies in the design of ConVERGD, namely AAV-based reporter technology and inspiration from the world of ribozymes, strands of RNA that can act like enzymes, catalyzing biochemical reactions.
Importantly, ribozymes can be engineered to control the on/off switch for gene expression with extreme precision.
“We originally heard about ribozymes from a journalism club that was thinking more therapeutically about how to use AAVs,” Schwartz said. “Alex came back and thought he could figure out a way to use them in neuroscience tools.”
Exciting for the neuroscience community and beyond
As a proof of concept, Schwartz and Hughes used ConVERGD to interrogate a subpopulation of norepinephrine neurons.
“Collectively, norepinephrine neurons do many different things,” Schwartz explained.
“The subset we targeted produced norepinephrine, but they also produced this other opioid peptide called dynorphin, which had not been characterized in these neurons before. With ConVERGD, we found that activation of only those dynorphin-expressing neurons was sufficient to elicit an anxiety response.
By analyzing the functions and assigning them to a subpopulation of cells, Schwartz hopes that targeted therapy is possible.
“We treat anxiety and depression with drugs that target norepinephrine signaling, but they target it globally,” Schwartz said.
“You’re also going to see impairment of other important functions for norepinephrine that you don’t want to see.” Targeting these neurons more specifically could help improve this.”
The work will have a ripple effect beyond St. Jude. “We’re really excited about this for the community,” Schwartz said. “ConVERGD should be responsive to any tissue. It may be useful beyond neuroscience.
Authors and funding
Other study authors include Brittany Pittman, Basie Xu, Jesse Gammons, Charisse Webb, Hunter Nolen, Philip Chapman and Jay Bikoff, St. Jude.
The study was supported by grants from the Brain and Behavior Research Foundation (NARSAD Young Investigator Grant), the National Institutes of Health (1DP2NS115764, P30 CA021765), and ALSAC, the fundraising and awareness organization of St. Jude.
About this neurotech research news
Author: Chelsea Bryant
source: St. Jude Children’s Research Hospital
Contact: Chelsea Bryant – St. Jude Children’s Research Hospital
Image: Image credited to Neuroscience News
Original research: Closed access.
“A single-vector cross-sectional AAV strategy to study cellular diversity and brain function” by Lindsay Schwarz et al. Nature Neurology
Summary
A single-vector cross-sectional AAV strategy for interrogating cellular diversity and brain function
As the discovery of cellular diversity in the brain accelerates, so does the need for tools to target cells based on multiple characteristics.
Here, we developed conditional viral expression by ribozyme-guided degradation (ConVERGD), an adeno-associated virus-based, single construct, cross-sectional targeting strategy that combines a self-cleaving ribozyme with traditional FLEx switches to deliver molecular cargo to specific neuronal subtypes.
ConVERGD offers advantages over existing cross-section expression platforms, such as extended cross-section targeting with up to five recombinase-based functions, accommodation of larger and more complex payloads, and a vector that is easy to modify for rapid toolbox expansion.
In the present report, we used ConVERGD to characterize an unexplored subpopulation of norepinephrine (NE)-producing neurons in the locus coeruleus of rodents that coexpress the endogenous opioid gene prodynorphin (Pdyn).
These studies demonstrate ConVERGD as a versatile targeting tool for a variety of cell types and reveal Pdyn-expression of NO+ locus coeruleus neurons as a small neuronal subpopulation capable of driving anxiogenic behavioral responses in rodents.
