The molecular reasoning behind neuronal diversity

summary: Researchers have explored how the types of nerve cells in the fruit fly fruit fly differentiate their functions even though they originate from a similar genetic framework.

In the study, two closely related subtypes of neurons expressed more than 800 different genes (about 5% of the fly’s genome) differently. This gene expression directly affected the differences observed between neuronal cell types.

The findings help shed light on the development of complex brain cells and how disease can affect them.

Key facts:

1. Two subtypes of related neurons in the fruit fly Drosophila showed differential expression of more than 800 genes, influencing their distinct functions.
2. Differences in gene expression helped explain functional differences such as the robust bursts of neurotransmitter release of phasic neurons compared to the more steady release of tonic neurons.
3. Technologies such as the isoform patchseq have enabled in-depth examination of gene expression, RNA editing, and splicing in individual neurons.

source: Picauer Institute for Learning and Memory

Discovering how hundreds of different types of brain cells develop through their unique expression of thousands of genes holds promise not only for advancing understanding of how the brain works in health, but also what goes on in disease.

A new MIT study thoroughly explores this “molecular logic” in two types of neurons Drosophila In fruit flies, even similar cells are shown to push and pull many levers to develop distinct functions.

In the study in nervous cellsA team of neurobiologists at the Picauer Institute for Learning and Memory found that the two closely related neural subtypes differ from each other in how more than 800 genes, or ~5% of the total genes encoded in the fly genome, are expressed.

By manipulating genes whose expression varies significantly, the scientists were then able to show how many of the remarkable differences between cells are produced.

“There is a global effort in neuroscience to identify all different types of neurons to determine their unique characteristics and gene expression profiles,” said study senior author Troy Littleton, a professor of neuroscience in MIT’s departments of Biology, Brain, and Cognitive Sciences.

“This information can be used as a toolkit to study how newly discovered disease genes relate to those specific neurons to indicate which cells may be most affected by specific brain disorders.”

We wanted to use Drosophila As a way to see if we can, in fact, determine how to use the transcriptome of two similar neurons differentially to understand the key genes that determine their unique structural and functional characteristics.

Under the microscope

The two neurons compared in the study emerge from the fly’s counterpart to the spinal cord to control muscles by releasing the neurotransmitter glutamate at connections called synapses.

The main functional differences between neurons are that “phasic” neurons connect to many muscles and emit large, episodic bursts of glutamate while each “excitatory” neuron connects to only one muscle and provides a more continuous drip of the chemical. This duality, also present in neurons in the human brain, provides a flexible range of control.

Suresh Geeti, a postdoctoral researcher at the Picauer Institute, led the effort in Littleton’s lab to determine how these two neurons developed their differences. The team started with an extraordinarily in-depth characterization of how the two types of cells differ in shape and function, then took a very precise look at their gene expression, or transcription profiles.

Upon closer examination, tonic and metaphase cells showed a variety of important differences. Physic neurons make fewer synapses on an individual muscle than do tonic cells, but because they innervate more muscles, phasic neurons have to make about four times as many synapses in total. Tonic neurons have more inputs than other neurons thanks to their wider-reaching dendrites (branches that lead into the cell).

On the output side of things, phasic neurons produced much stronger signals when stimulated and were more likely to transmit them than tonic neurons. The analysis showed that the synaptic sites that stimulate glutamate release, called activated zones (AZs), receive more calcium ions in phase neurons than in tonic neurons.

A particularly interesting new finding was that the AZ regions in tonic and phase neurons took on different shapes. The thousand tonic regions were round, like donuts, while the phase ones were more triangular or star-shaped. Littleton hypothesizes that this difference could allow more calcium ions to collect in the active phase regions, possibly explaining their greater bursts of glutamate release than in tonic neurons.

express their differences

To assess gene expression, Getty used a technique called isoform patchseq in which he identified the same tonic and phasing neurons in hundreds of flies and extracted RNA from their individual nuclei and cell bodies.

The technique, while extremely hard work, Littleton said, provided the team with an unusually rich source of transcriptional information from specifically the cells of interest, including not only how gene expression differs between the two cell types, but also how genes are linked. and RNA editing. She was different.

Overall, the expression of 822 genes was significantly different between the two types of neurons. About 35 genes are known to help direct the development of axonal branches that neurons extend to form their connections with muscles—a set of differences related to why tonic neurons innervate only one muscle while phasic neurons innervate many.

Other differentially expressed genes related to the structure and function of synapses, while more than 20 others suggested differences in the neuromodulatory chemicals each neuron was sensitive to as inputs.

The team found that transport proteins are more expressed in metaphase neurons, possibly explaining how they keep up with the greater demand for more synapses across many muscles. The team also found that while tonic neurons express silyl genes to bind sugars to proteins on their synaptic membrane, tonic neurons express unique ubiquitin genes that break down the proteins.

After documenting the most notably different genes, the team set out to determine what they do by disrupting their function and see how this affects cells.

For example, Getty, Littleton and colleagues found that interference with specific ubiquitin genes caused an increase in the growth of metaphasic neurons at synapses. Meanwhile, disruption of the sialolysis process decreased synaptic growth in the tonic neurons. The tonic neurons also expressed 40 times more of a gene called Wnt4, and inactivation of Wnt4 reduced synaptic growth in this group of neurons.

The scientists also found that phasic neurons express a calcium buffer gene more than 30 times more than tonic genes. When they mutated this gene to disable its function, they found that phasic neurons, which normally have low basal levels of calcium, now display higher resting calcium similar to tonic neurons.

In another experiment, they showed that they could explicitly disrupt the AZ forms of each cell by interfering with the cytoskeletal genes that each neuron specifically expressed. When the team reduced a gene that metastatic neurons express a lot, their AZ regions became elongated, but their activated AZ regions were unaffected. When the team significantly reduced a gene expressed by metaphasic neurons, their AZ regions became less round without affecting the AZ regions of metaphasic cells.

Overall, the analysis enabled the team to begin to build a model of the molecular differences that make the two cells different, though Littleton said they still have more work to do to understand how the full range of gene expression differences determine the unique characteristics of the two neurons. Subspecies.

In addition to Littleton and Getty, other authors of the paper are Andres Crane, Yulia Akbarjinova, Nicole Aponte Santiago, Karen Cunningham, and Charles Whitaker.

The JPB Foundation, the Picquer Institute for Learning and Memory, and the National Institutes of Health funded the research.

About genetics and neuroscience research news

author: David Ornstein
source: Picauer Institute for Learning and Memory
communication: David Ornstein – Picauer Institute for Learning and Memory
picture: Image credited to Neuroscience News

Original search: open access.
“Molecular logic of synaptic diversity between Drosophila tonic and phasic motoneuronsWritten by Troy Littleton et al. nervous cells

a summary

Molecular logic of synaptic diversity between Drosophila tonic and phasic motoneurons

Highlights

• Motor and excitatory neurons show a diversity of synaptic structure and output
• Isoform-Patch-seq highlights differentially regulated molecular pathways
• Genetic analyzes identify regulators of Ca²⁺ Caching and organizing AZ
• Differential ubiquitination and cellulose regulate synapse growth and structure

summary

Although neuronal subtypes display unique synaptic organization and function, the underlying transcriptional differences that establish these features are poorly understood. To identify the molecular pathways that contribute to synaptic diversity, Patch-seq RNA patterning of single neurons was performed on Drosophila Tonic motor neurons and the glutamatergic phase.

PT neurons form weaker synapses on single muscles, while PT neurons form stronger synapses on multiple muscles. Ultra-resolution microscopy and in vivo Imaging showed that active synapses in phasic motor neurons are tighter and display enhanced calcium exposure²⁺ flow compared to tonic counterparts.

Genetic analysis identified unique synaptic properties mapping to gene expression differences of several cellular pathways, including distinct signaling ligands, posttranslational modifications, and intracellular calcium.²⁺ Stores.

These findings provide insight into how unique transcriptomes drive functional and morphological differences between neuronal subtypes.