Immune Proteins Play a Role in Neural Growth

Post by Anastasia Sares

What's the science?

The protein IL-33 is part of a large family of proteins related to immune function. Outside of the brain, it is mostly known for its role in inflammation, such as in asthma or auto-immune conditions. However, IL-33 also supports tissue repair and is necessary for proper neural development. In studies of brain injury and Alzheimer’s disease, IL-33 has been shown to increase plasticity and reverse memory deficits. But how can an immune system protein support memory? This week in Cell, Nguyen and colleagues examined IL-33’s role in memory formation in the hippocampus and showed that it involves the brain’s immune cells to clear a path for branching neurons. 

How did they do it?

The research team employed a number of techniques for identifying cells producing IL-33 or its receptor, IL1RL1, in genetically modified mice. Some of the mice had a code for a fluorescent protein inserted right next to the IL-33 gene so that cells producing IL-33 could be seen with a fluorescent microscope. Others had genetic manipulations to knock out the IL-33 gene so that it no longer worked. The authors were also able to express extra IL-33 in mice using a lentivirus.

Their analyses of brain tissue included flow cytometry, which involves streaming a solution full of cell nuclei in a small tube one cell wide. As the nuclei passed one by one in front of a laser, they could measure many different properties of the scattered laser light to detect different proteins (including the fluorescence from the IL33 gene). The authors also raised mice in different conditions. Some of them experienced temporarily enriched environments, known for increasing neuronal growth and plasticity, while others experienced brief social isolation, which does the opposite. Still, others underwent fear conditioning to see how well they learned the association between a stimulus and a mild shock. 

What did they find?

In the hippocampus, IL-33 was expressed primarily in neurons (and especially neurons in the dentate gyrus). This is different from elsewhere in the brain, where it is mostly found in astrocytes. The receptor, IL1RL1, was located in microglia, which are cells acting as the primary form of immune defense in the central nervous system.

IL-33 levels increased in the mice who had enriched environments and decreased in those undergoing social isolation. IL-33 also decreased with age and memory loss. Deleting IL-33 or its receptors on the microglia caused mice to have fewer dendritic spines on existing neurons, fewer new neurons, and less precise memory for fearful stimuli. Extensive testing indicated that IL-33 might induce the microglia to clear out the extracellular matrix (the web of structural proteins around the cells). This clearance would then pave the way for the growth and branching of new neurons. 

What's the impact?

IL-33 may have neuroprotective benefits in diseases like stroke, traumatic brain injury, ALS, and even Alzheimer’s disease (which specifically affects memory and could be related to inflammation in the brain). Understanding the mechanisms behind synapse development and memory maintenance will help us to fight these diseases. This research also emphasizes and expands the role of immune cells like microglia in brain function.

Nguyen et al. Microglial Remodeling of the Extracellular Matrix Promotes Synapse Plasticity. Cell (2020). Access the original scientific publication here.

Converting Astrocytes Into Neurons Reverses Motor Deficits in a Model of Parkinson’s Disease

Post by Amanda McFarlan

What's the science?

Parkinson’s disease is a neurodegenerative disorder associated with motor deficits caused by the substantial loss of dopaminergic neurons in the substantia nigra (a region involved in reward and movement). Recent studies have shown that it is possible to reprogram fibroblasts (cells involved in synthesizing the extracellular matrix and collagen) to become neurons by downregulating the RNA-binding proteins polypyrimidine tract-binding protein (PTB) and nPTB (another member of the PTB family) since both PTB and nPTB suppress factors that prevent cells from developing into a neuron. Like fibroblasts, glial cells in the brain known as astrocytes are also flexible with respect to cell fate, however, whether they can be converted to neurons is still unknown. This week in Nature, Qian and colleagues investigated whether astrocytes in the substantia nigra could be converted into functional dopaminergic neurons and whether this conversion could restore neuronal populations and motor deficits in a chemically-induced model of Parkinson’s disease.

How did they do it?

To determine whether astrocytes could be converted into neurons, the authors used reverse transcription qPCR to measure the expression levels of neuron-specific transcription factors and microRNAs in mouse astrocytes from the cortex and midbrain. They hypothesized that, unlike fibroblasts, the knockdown of PTB alone would be sufficient to convert astrocytes into neurons. To test this hypothesis, the authors disrupted PTB signalling in-vitro in cell cultures by transducing astrocytes from both mouse and human tissue with a virus containing a small hairpin RNA against PTB. Eight weeks later, the authors determined whether the astrocytes had been converted to neurons by using patch clamp recordings to assess their electrophysiological properties. Then, the authors investigated whether astrocytes could be converted into neurons in vivo by using viral injections to induce PTB knockdown in astrocytes in the mouse midbrain. Ten weeks after the viral injections, the authors confirmed whether the astrocytes in the substantia nigra of the midbrain had been successfully converted into dopaminergic neurons by assessing their morphological properties and expression of neuronal markers. They then investigated whether the conversion of astrocytes into dopaminergic neurons in the substantia nigra could be used to reconstitute the neuronal population in this area following an injury. They used the 6-hydroxydopamine (a neurotoxin that destroys dopaminergic neurons) model of Parkinson’s disease to deplete the substantia nigra of dopamine neurons and then performed viral injections to induce PTB knockdown in astrocytes as previously described. Finally, the authors used chemogenetics and behavioural tests to determine whether injury-induced impairments to mouse motor function can be restored by converting astrocytes in the injured area into neurons.

What did they find?

The authors determined that up to 80% of astrocytes displayed neuronal morphologies only four weeks after in-vitro viral transduction. These cells also exhibited neuron-specific electrophysiological properties including voltage-gated sodium and potassium channels and action potential firing, suggesting that PTB knockdown is sufficient to convert astrocytes into neurons in both mouse and human tissue. Then, the authors revealed that the in vivo conversion of astrocytes to neurons in the substantia nigra was a gradual process, with 20% conversion at three weeks, 60% conversion at five weeks, and finally, 80% conversion at ten weeks post-injection. Moreover, they observed that these dopaminergic neurons projected to areas of the nigrostriatal pathway including the caudate, putamen, and the nucleus accumbens. Together, these findings suggest that astrocytes in the substantia nigra can be converted into dopamine neurons that are re-incorporated into the nigrostriatal pathway in a time-dependent manner.

Next, the authors showed that the conversion of astrocytes to neurons restored the population of dopaminergic neurons in the substantia nigra to 33% of the initial number prior to injury. Injury-induced motor deficits resulting from the depletion of dopaminergic neurons were gradually restored to almost normal levels following the viral transduction of astrocytes. Furthermore, chemogenetic manipulations confirmed that these improvements in motor deficits were due to the reconstitution of the dopaminergic neuronal population in the substantia nigra. 

What’s the impact?

This is the first study to show that it is possible to reprogram astrocytes in the substantia nigra of the mouse brain to become functional dopaminergic neurons. Moreover, the authors demonstrated that this conversion of astrocyte to neuron could be used to partially restore the population of dopaminergic neurons in the substantia nigra as well as improve motor deficits following a chemically induced lesion to this area. Together, these findings provide exciting new insights into possible treatments that could be developed to treat individuals with neurodegenerative disorders, like Parkinson’s disease, that are associated with a substantial loss of neuronal populations. 

Qian et al. Reversing a model of Parkinson’s disease with in situ converted nigral neurons. Nature (2020). Access the original scientific publication here.

Effects of Neurofeedback on Emotional Regulation in Adolescent Females

Post by Elisa Guma

What's the science?

Adolescence is characterized by a wide range of neural changes that subserve higher order socio-emotional and cognitive function. During this transition period deficits in emotional regulation may emerge putting individuals at higher risk for developing psychiatric illnesses. The structural and functional connectivity of the amygdala and prefrontal cortex (PFC) are thought to play a key role in modulating emotional regulation, however, it is unclear whether we can intervene to actively shape these connectivity patterns to improve emotional and cognitive abilities during this critical time. This week in Neuroimage, Zich and colleagues aim to investigate if they can successfully use neurofeedback (a type of biofeedback aimed at teaching individuals self-control of brain activity via visual or audio feedback) from functional magnetic resonance imaging (fMRI) to help individuals regulate their emotions in real time.

How did they do it?

The authors conducted three different experiments to investigate whether neurofeedback based on an individual’s own PFC-amygdala connectivity could be used to modulate neural and emotional measures relevant for emotion regulation. In all experiments, adolescent females were evaluated on their emotional regulation abilities, as well as a variety of mood and anxiety measures. The first experiment was used to determine the best neurofeedback implementation. During the neurofeedback task participants were shown an image of a ten-segment thermometer while undergoing fMRI. During blocks in which no neurofeedback was occurring, the temperature was frozen at 6/10, whereas during neurofeedback blocks, the temperature of the thermometer was a direct real-time reflection of the PFC-amygdala connectivity. Participants were asked to try to control the temperature of the thermometer by controlling their thoughts and feelings and revisiting emotional reappraisal strategies. The three different implementations varied slightly in the way the thermometer displayed a change in connectivity.

Experiment 2 replicated experiment 1, except with a larger sample, and the best neurofeedback implementation (gleaned from experiment 1). Finally, in Experiment 3, the number of neurofeedback blocks was doubled, and the authors also collected Magnetic Resonance Spectroscopy data from the anterior cingulate cortex (implicated in reward processing) and PFC to measure Gamma aminobutyric acid (GABA) and glutamate, the major inhibitory and excitatory neurotransmitters in the brain. 

What did they find?

In their first experiment, the authors identified the best neurofeedback implementation and found a negative reinforcement of functional connectivity to be optimal (i.e. if the thermometer changes from 2/10 to 3/10 there was a more negative correlation of PFC-amygdala connectivity). In experiment 2, the authors aimed to assess the effects of one neurofeedback session on neural, emotional/cognitive measures, and their association in a larger sample. They did not find a significant change in emotional/cognitive measures or in the change in functional connectivity at the group level, but they did observe a practice-related change in connectivity, related to changes in thought control ability. Further, they found that state anxiety before the MRI session influenced the difference between functional connectivity in neurofeedback blocks relative to those with no neurofeedback.

Finally, in experiment 3 the authors replicated the findings from above in an independent sample. Moreover, they found that the concentrations of the inhibitory neurotransmitter, GABA ⁠— in both the PFC and anterior cingulate cortex ⁠— moderated the relationship between state anxiety before the MRI session and the effect of neurofeedback on PFC-amygdala functional connectivity.

What's the impact?

The study provides evidence for the feasibility of using neurofeedback in adolescent females to modulate functional connectivity measures between the prefrontal cortex and the amygdala. Further, the authors show that the relationship between state anxiety and the effect of the neurofeedback was modulated by GABA concentrations in the PFC and anterior cingulate cortex. Future studies may investigate the effects of longer training sessions and extend this work into populations with neuropsychiatric disorders.

Zich et al. Modulatory effects of dynamic fMRI-based neurofeedback on emotion regulation networks in adolescent females. Neuroimage (2020). Access the original scientific publication here