A revolutionary tool reveals the hidden functions of brain cells

Abstract: Researchers have developed ConVERGD, a tool for precise manipulation of specific cell subpopulations, improving studies of cell diversity. The study demonstrated the utility of ConVERGD by identifying a subpopulation of norepinephrine neurons associated with anxiety. This innovative approach could impact research and treatment in a variety of fields.

Key facts:

• A precise tool: ConVERGD enables precise targeting and manipulation of cell subpopulations.
• Application in neuroscience: Used to identify the norepinephrine neuron subpopulation that influences anxiety.
• Broad impact: Potential applications beyond neuroscience, benefiting different areas of research.

Source: Children’s Research Hospital St. Judas

As gene sequencing technologies become more powerful, our understanding of cellular diversity grows in parallel. This led scientists from the Children’s Research Hospital St. They hope to create a tool to improve the ease and accuracy 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 intersectional expression platforms by accommodating more complex genetic load and increased adaptability.

The researchers demonstrated the utility of ConVERGD by studying a previously unidentified subpopulation of norepinephrine neurons. The paper demonstrates the significant impact that research into cell subpopulations could have on basic research and health care.

The findings were published today in The neuroscience of nature.

Same type of cells, different functions

For Dr. Lindsay Schwarz, St. Jude Department of Developmental Neurobiology, necessity prompted the invention as she explored the landscape of neuronal cells, specifically neurons that produce norepinephrine.

“Norepinephrine neurons are thought to be just one type of neuron. But when they’re activated in the brain, they can cause a lot of different kinds of behavior, like increasing attention and memory formation or causing a stress response or a fight-or-flight response,” Schwarz said.

“But if only one type of neuron releases this one molecule, then how does that make you do different things?”

Investigating such questions requires the ability to selectively probe cell subpopulations with extreme biases. To that end, Schwarz found that all attempts to use current practices failed.

“We didn’t go into this project thinking we were going to build a new tool, but it felt like a need in the community.”

Improving current cell subpopulation targeting technology

Targeting subpopulations of cells requires passing them through several genetic filters. These intersection filters examine which genes cells express and which pathways and connections they make, analyzing different subpopulations so 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 to specific cells with high precision is an ideal approach for the application of these intersection filters.

These reporter tools are used to label or monitor gene expression and protein localization within specific cells or regions. However, they can be complex to design and offer a limited amount of space inside.

“One of our main goals was to design a tool where your gene of interest is only expressed when it is carried with multiple features, but can be easily modified by end users to put in whatever genes they want,” Schwarz explained.

Robust ribozymes offer next-generation specificity

Schwarz and the first author, Ph.D. Alex Hughes, graduate of St. Jude Graduate School of Biomedical Sciences, now the Allen Institute for Brain Science, used two separate technologies in the design of ConVERGD, namely an AAV-based reporter technology and inspiration from the world of ribozymes, RNA chains that can act like enzymes catalyzing biochemical reactions .

Importantly, ribozymes can be engineered to control the on/off switches of gene expression with extreme precision.

“We first heard about ribozymes from a journal club that was thinking more about how to use AAV therapeutically,” Schwarz said. “Alex came back and figured he could figure out a way to use them in neuroscience tools.”

Exciting for the neuroscience community and beyond

As a proof of concept, Schwarz and Hughes used ConVERGD to examine a subpopulation of noradrenaline neurons.

“Together, norepinephrine neurons do a lot of different things,” Schwarz explained.

“The subset we targeted produce norepinephrine, but they also produce this other opioid peptide called dynorphin, which has not been previously characterized in these neurons. With ConVERGD, we found that activating just these dynorphin-expressing neurons is enough to cause an anxiety response.”

By dissecting the functions and assigning them to subpopulations of cells, Schwarz hopes there is a possibility of targeted therapy.

“We treat anxiety and depression with drugs that target noradrenaline signaling, but they target it globally,” Schwarz said.

“You’ll also see damage to other important functions of 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. Judas. “We’re really excited about this for the community,” Schwarz said. “ConVERGD should be amenable to every tissue. It could be useful outside of neuroscience.”

Authors and funding

Other study authors include Brittany Pittman, Beisi Xu, Jesse Gammons, Charis Webb, Hunter Nolen, Phillip Chapman and Jay Bikoff, St. Judas.

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 St. Jude to raise funds and awareness.

About this neurotechnology research news

Author: Chelsea Bryant
Source: Children’s Research Hospital St. Judas
Contact: Chelsea Bryant – Children’s Research Hospital St. Judas
Picture: Image credited to Neuroscience News

Original research: Closed access.
“A single-vector intersectional AAV strategy for probing brain cell diversity and function” Lindsay Schwarz et al. The neuroscience of nature

Abstract

A single-vector intersectional AAV strategy for probing brain cell diversity and function

As the discovery of cellular diversity in the brain accelerates, so does the need for tools that target cells based on multiple features.

Here, we developed conditional viral expression by ribozyme-guided degradation (ConVERGD), an adeno-associated virus-based intersectional targeting strategy, a single construct, that combines a self-cleaving ribozyme with traditional FLEx switches to deliver molecular cargo to specific neuronal subtypes.

ConVERGD offers advantages over existing intersectional expression platforms, such as extended intersectional targeting with up to five recombinase-based features, accommodation of larger and more complex payloads, and an easy-to-modify vector for rapid expansion of the toolkit.

In this report, we used ConVERGD to characterize an unexplored subpopulation of norepinephrine (NE)-producing neurons within the locus coeruleus of rodents coexpressing the endogenous opioid gene prodynorphin (Pdyn).

These studies demonstrate ConVERGD as a versatile tool for targeting different cell types and reveal Pdyn– expression of NO⁺ locus coeruleus neurons as a small neuronal subpopulation capable of triggering anxiogenic behavioral reactions in rodents.