Imaging Technique Finds Key Neurons in Brain Interact, May Support More Targeted Treatments

nerve cell communication

Two key types of brain nerves cells affected by Parkinson’s disease — cholinergic neurons and dopaminergic neurons — communicate and interact via signaling systems, researchers were able to “see” using a new imaging approach.

This beneficial neuron-to-neuron interaction, confirmed through the novel approach in a rat model of the disease, also supported further work on targeted treatments for Parkinson’s, including a potential gene therapy.

Their study, DREADD Activation of Pedunculopontine Cholinergic Neurons Reverses Motor Deficits and Restores Striatal Dopamine Signaling in Parkinsonian Rats,” was published in Neurotherapeutics.

Parkinson’s is a progressive neurodegenerative disease, meaning that it steadily worsens as neurons die over time. One of its hallmarks is the loss of dopamine — a neurotransmitter crucial for coordinating movement and regulating mood — that occurs when dopaminergic neurons in a brain structure called the substantia nigra malfunction and die.

Cholinergic neurons — those that produce the neurotransmitter acetylcholine — are nerve cells found in the pedunculopontine nucleus (PPN) of the brain. They are also implicated in Parkinson’s, since in post mortem studies of patients’ brain tissue a significant amount of these cells are found dead.

Researchers had previously used used a harmless virus to deliver a genetic modification to cholinergic neurons in a rat model of Parkinson’s disease. This technique is called designer receptors exclusively activated by designer drugs (DREADDs), and consists of a class of engineered proteins that allow researchers to hijack cell signaling pathways in order to look at cell-to-cell interactions more easily.

The animals were then given a compound designed to activate the genetic ‘switch’ and stimulate the target neurons. After treatment, almost all animals had recovered and were able to move.

Now, this same research team used positron emission tomography (PET), a brain imaging technology, together with DREADDs to selectively activate cholinergic neurons in the brains of diseased rats and look at how other brain cells responded.

They found that stimulating cholinergic neurons led to the activation of dopaminergic neurons in the rat brain, and dopamine was released.

This means that cholinergic activation restored the damaged dopaminergic neurons. The parkinsonian rats appeared to completely recover — they were able to move without problems and their postures returned to normal.

“This is really important as it reveals more about how nerve systems in the brain interact, but also that we can successfully target two major systems which are affected by Parkinson’s disease, in a more precise manner,” Ilse Pienaar, PhD, a researcher at the University of Sussex and Imperial College London and study author, said in a press release.

“While this sort of gene therapy still needs to be tested on humans, our work can provide a solid platform for future bioengineering projects,” Pienaar added.

This new technique has several advantages over deep brain stimulation (DBS), a surgical procedure that sends electrical impulses to the brain to activate the neurons.

Deep brain stimulation can help to relieve some Parkinson’s symptoms, but is invasive and has had mixed results. Some patients show improvements while others experience no changes in symptoms or even a deterioration. This may be due to therapy imprecision, as DBS stimulates all types of brain nerve cells without a specific target.

This study sought to address the selectivity issue by looking at the activation of one type of cell in a specific part of the brain to get a better understanding of how other parts might be influenced.

“[T]he current data could allow for designing medical approaches capable of improving the ratio between desirable and undesirable outcomes and leaving nonimpaired functions intact. For example, specific genetically defined neurons … could be targeted to treat motor symptoms of [Parkinson’s], without inducing a cognitive detriment, and vice versa,” the researchers wrote.

“For the highest chance of recovery, treatments need to be focused and targeted but that requires a lot more research and understanding of exactly how Parkinson’s operates and how our nerve systems work,” Pienaar said. “Discovering that both cholinergic and dopaminergic neurons can be successfully targeted together is a big step forward.”

The researchers concluded, “[t]his study supports the hypothesis that it is the cholinergic neuronal population, projecting from the PPN, which delivers some of the clinical benefits associated with PPN-DBS.”

Pienaar and colleagues collaborated with Invicro, a precision medicine company, for this study. Lisa Wells, PhD, a study co-author on the study and Invicro employee added, “It has been an exciting journey … to combine the two technologies [DREADD and PET] to offer us a powerful molecular approach to modify neuronal signaling and measure neurotransmitter release. We can support the clinical translation of this ‘molecular switch’ … through live imaging technology.”

This work may make possible more selective and more effective treatment alternatives to deep brain stimulation.

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Number of Dopaminergic Neurons at Birth Could Affect Lifetime Parkinson’s Risk, Report Suggests

Dopaminergic neurons at birth

The number of dopamine-producing (dopaminergic) neurons at birth could influence a person’s lifetime risk of developing Parkinson’s disease, according to a group of experts.

Because these nerve cells die over the course of the disease, having fewer of them to begin with could translate to a higher risk, the scientists said.

Their report, “Does Developmental Variability in the Number of Midbrain Dopamine Neurons Affect Individual Risk for Sporadic Parkinson’s Disease?” is a review of scientific literature on the topic. It was published in the Journal of Parkinson’s Disease.

Parkinson’s is a progressive neurodegenerative disease, meaning that it steadily worsens as neurons die over time. The hallmark of Parkinson’s is the loss of dopamine — a neurotransmitter crucial for coordinating movement and regulating mood — that occurs when dopaminergic neurons in a brain structure called the substantia nigra malfunction and die.

The substantia nigra communicates with a neighboring brain structure called the striatum and it is believed that the loss of dopamine and dopaminergic neurons in these structures must cross a certain threshold before the symptoms of Parkinson’s become noticeable. The number of dopaminergic neurons an individual is born with, therefore, might influence how soon this threshold is reached.

Precisely how many of these neurons must die before symptoms appear remains an open question. No datasets of nigral (belonging to the substantia nigra) dopaminergic neuron counts are available for individuals with recent onset of Parkinson’s symptoms.

Nor is there a strong scientific consensus regarding the number of dopaminergic neurons in normal substantia nigra. Different methods for marking both dopaminergic neurons and age-related changes that can limit the number of functional neurons within the brain hinder precise counts.

To estimate the variation in dopaminergic neuron numbers across people, a team of scientists now examined the data in four previous studies. Each study followed strict exclusion criteria, such as not admitting patients with histories of neuropsychiatric disease and/or other neurological damage.

The studies are: Ageing of substantia nigra in humans: cell loss may be compensated by hypertrophy, published in 2002; The absolute number of nerve cells in substantia nigra in normal subjects and in patients with Parkinson’s disease estimated with an unbiased stereological method, published in 1991; Unbiased morphometrical measurements show loss of pigmented nigral neurones with ageing, published in 2002; and Morphometry of the human substantia nigra in ageing and Parkinson’s disease, published in 2008.

In the review, the researchers focused on individuals who died before their 51st birthday, to minimize the risk that any observed variation was due to age-related effects, or to undiagnosed progressive disorders.

The studies showed a wide variation between individuals — ranging from 147% to 433% — in terms of the difference between those with the most and the fewest dopaminergic neurons. Such variation must be better understood to properly understand its significance, the researchers said.

Many of the genes implicated in rare developmental abnormalities in humans also are involved in determining the location, formation, and size of the dopaminergic neuron population. Based on past studies, the researchers suggest that subtle changes in how active these genes are throughout development likely determine the variation witnessed between individuals.

Although clearly defining these changes poses a significant challenge, some clues are emerging.

One recent study, for example, linked Parkinson’s risk to single nucleotide polymorphisms (SNPs) — changes of one single letter of the genetic code — that affected gene regulatory elements involved in early nervous system development. This, in turn, affects the number of neurons an organism has.

Other studies have shown that Parkinson’s-related mutations also can reduce the number of dopaminergic neurons capable of being grown in the lab.

The alpha-synuclein protein plays a well-documented role in Parkinson’s progression. Its role in early development is less understood. One mouse study showed that the expression of the alpha-synuclein gene could affect the number of dopaminergic neurons in the substantia nigra. This suggests that, beyond the pathogenic role it plays in Parkinson’s, alpha-synuclein may help determine the early development and survival of dopaminergic neurons.

Non-genetic factors also appear to impact dopaminergic neurons by affecting critical periods of brain development. These factors include viral infection, exposure to environmental toxins, and hypoxia (low oxygen) at birth.

Based on the information collected throughout their review, the researchers propose that the number of nigral dopaminergic neurons individuals are born with and that survive immediately following birth affects their lifetime risk of developing Parkinson’s disease.

However, the researchers note that current knowledge of the factors influencing the development and survival of dopaminergic neurons is incomplete.

Therefore, “we need to explore the changes that occur both during development and during adulthood and aging when we seek to understand the full landscape of [Parkinson’s] risk,” they said.

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Awakening Dormant Neurons Could Provide Disease-modifying Parkinson’s Treatment, Early Study Suggests

dormant neurons

Together with dying nerve cells, dormant neurons also may be at the root cause of Parkinson’s disease, according to a recent study in animal models.

Reawakening these neurons by targeting a type of brain cells called astrocytes can restore dopamine production in the brain and reverse Parkinson’s motor symptoms, the study found. These findings could lead to a potential new disease-modifying treatment, especially at the early stages of Parkinson’s.

The study, “Aberrant Tonic Inhibition of Dopaminergic Neuronal Activity Causes Motor Symptoms in Animal Models of Parkinson’s Disease,” was published in the journal Current Biology.

Despite its prevalence and debilitating consequences, current medical therapy for Parkinson’s relies on alleviating symptoms. Research investigating ways of modifying the disease or reversing its symptoms is scarce, based on the firm belief that Parkinson’s is caused by the irreversible death of nerve cells — also called neurons — in a region of the brain called the substantia nigra.

In this brain region, nerve cells known as dopaminergic neurons are responsible for producing the neurotransmitter dopamine, a chemical messenger that allows nerve cells to communicate. Dopamine plays a key role in motor function control and also is involved in behavior and cognition, memory and learning, sleep, and mood.

Levodopa, a mainstay of Parkinson’s treatment, works by supplying extra dopamine to the brain. However, it only alleviates motor symptoms and does not alter the disease course. Moreover, its long-term use can cause serious side effects, including involuntary, erratic, and writhing movements.

Now, a team of Korean researchers have discovered additional clues about the underlying mechanisms of Parkinson’s that may offer hope for the development of disease-modifying treatments that could reverse the condition.

Using mouse and rat models of Parkinson’s, they found that the motor abnormalities that mark the disease begin earlier than was previously thought. They are triggered when dopaminergic neurons in the substantia nigra are still alive but in a dormant state, unable to produce dopamine.

However, what holds the key to that dormant state is another type of cells called astrocytes, star-shaped cells present in the brain and spinal cord that play important roles in the protection and regulation of the nervous system.

When neurons die, nearby astrocytes react by proliferating, and start to release an inhibitory neurotransmitter called gamma-aminobutyric acid (GABA) at excessive levels. This puts neighboring neurons “on hold,” suspending their production of dopamine.

GABA prevents the neurons from firing electrical impulses and causes them to stop making an enzyme, called tyrosine hydroxylase, that’s essential for the production of dopamine. In effect, GABA puts the neurons into a dormant, or sleeping state.

One of the most important discoveries of the study was that surviving dormant neurons could actually be “awakened” from their “sleeping” state and rescued to alleviate motor symptoms.

“Everyone has been so trapped in the conventional idea of the neuronal death as the single cause of PD. That hampers efforts to investigate roles of other neuronal activities, such as surrounding astrocytes,” C. Justin Lee, PhD, the study’s corresponding author, said in a press release.

“The neuronal death ruled out any possibility to reverse PD. Since dormant neurons can be awakened to resume their production capability, this finding will allow us to give PD patients hopes to live a new life without PD,” Lee added.

Treatment with two different compounds that block GABA production in astrocytes, called monoamine oxidase-B, or MAO-B, inhibitors, was sufficient for neurons to recover the enzymatic machinery necessary to produce dopamine, the study found. This significantly alleviated Parkinson’s motor symptoms in the study animals.

In fact, the MAO-B inhibitors used for the study — selegiline (brand names EldeprylCarbex, Zelapar, among others), and safinamide (brand name Xadago) — are already prescribed to Parkinson’s patients as an add-on therapy to levodopa. They are believed to prevent the break down of dopamine in the brain.

Importantly, the existence of dormant neurons was observed in the brains of human patients. Analysis of postmortem brains of individuals with mild and severe Parkinson’s had a significant population of dormant neurons surrounded by numerous GABA-producing astrocytes.

The researchers hope that “awakening” neurons using MAO-B inhibition could be an effective disease-modifying therapeutic strategy for Parkinson’s, especially for patients in the early stages of the disease. At that time, inactive, yet live dopaminergic neurons are still present.

Although the results from several clinical trials have cast doubt on the therapeutic efficacy of traditional MAO-B inhibitors, researchers say they have recently developed a new inhibitor, KDS2010. KDS2010 effectively inhibits astrocytic GABA production with minimal side effects in Alzheimer’s animal models and also could be effective for alleviating Parkinson’s motor symptoms, the investigators said.

“This research refutes the common belief that there is no disease-modifying treatment for PD due to its basis on neuronal cell death,” said Hoon Ryu, PhD, a researcher at KIST Brain Science Institute, in South Korea, and one of the senior authors of the study.

“The significance of this study lies in its potential as the new form of treatment for patients in early stages of PD,” Ryu said.

The fact that inhibition of dopaminergic neurons by surrounding astrocytes is one of the core causes of Parkinson’s should be a “drastic turning point” in understanding and treating not only Parkinson’s but also other neurodegenerative diseases, added Sang Ryong Jeon, MD, PhD, also a researcher at KIST and a study co-author.

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Baicalin Protected Rats Against Parkinson’s Neurodegeneration

baicalin study

A bioactive agent called baicalin prevented neurodegeneration of Parkinson’s disease in rats by protecting against oxidative stress and neuronal death, according to a recent study.

The results, “Neuroprotective effect and mechanism of baicalin on Parkinson’s disease model induced by 6-OHDA,” were published recently in the journal Neuropsychiatric Disease and Treatment.

Although Parkinson’s trigger is unknown, research indicates its causative mechanism involves genetics, malfunction of mitochondria (the cells’ “powerhouses”), and oxidative stress — an imbalance between the production of harmful free radicals and the ability of cells to detoxify them, resulting in cellular damage.

Taken together, these molecular and cellular changes eventually result in the death of dopamine-producing neurons, the nerve cell type that is gradually lost in Parkinson’s disease.

Available treatments only ease disease symptoms, and there are currently no disease-modifying therapies that can delay or prevent Parkinson’s neurodegeneration.

Baicalin, a compound isolated from the Chinese skullcap‘s (Labiatae Scutellaria Linn Scutellaria baicalensis Georgi) dry roots, has been shown to have antibacterial, antiviral, anti-inflammatory, anti-tumor, cardiovascular, and neuroprotective activities.

Importantly, evidence shows that baicalin protects against dopaminergic neuronal damage induced by either rotenone or MPTP, two neurotoxins that are commonly used to replicate Parkinson’s in animal models.

A Chinese team of researchers now investigated the effects of baicalin on a 6-hydroxydopamine (6-OHDA)-induced rat model of Parkinson’s disease. Like rotenone and MPTP, 6-OHDA induces the death of dopamine-producing neurons and mimics Parkinson’s symptoms.

Baicalin was given in one of three doses: low (50 mg/kg), medium (100 mg/kg), or high (150 mg/kg). Following baicalin continuous administration for eight weeks, scientists assessed animals’ fatigue, motor coordination, voluntary movement, anxiety and exploratory behavior on a weekly basis. Neuronal changes following baicalin treatment also were evaluated.

Baicalin was found to improve rats’ coordination and voluntary movement. The compound also prevented oxidative stress-related neuronal damage and death, and promoted the release of neurotransmitters to regulate dopamine-dependent communication within the rats’ brain by regulating six small metabolic molecules: N-acetyl-aspartate (NAA), aspartate, glutamate, gamma-aminobutyric acid, glycine, and taurine.

“NAA is a hallmark of neuronal changes in the brain, and a decreased level suggests a loss or dysfunction of neurons,” researchers noted. On the other hand, glutamate is mainly involved in signal transmission, and learning and memory formation.

Further analysis revealed rats with Parkinson’s had low levels of N-acetyl-aspartate (NAA) and high levels of glutamate in the striatum (a brain region involved in motor control). After continuous administration of baicalin for two months, NAA and glutamate concentrations in the striatum changed in a dose-dependent manner to almost similar levels of those seen in healthy animals: higher baicalin doses resulted in increased metabolite concentrations.

Importantly, the team believes that both NAA and glutamate levels could be potential diagnostic biomarkers to assess neurodegeneration in the context of Parkinson’s disease.

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New Gene Therapy Delivery Method May Open BRAVE New World in Parkinson’s Research

BRAVE gene therapy method

A new method allows researchers to develop adeno-associated virus (AVV) — commonly used as the vehicle for gene therapies — that accurately target and deliver genes to specific cells in the body.

This new technology may be suitable to target dopaminergic neurons that are damaged in Parkinson’s disease.

“We believe that the new synthetic [lab-made] virus we succeeded in creating would be very well suited for gene therapy for Parkinson’s disease, for example, and we have high hopes that these virus vectors will be able to be put into clinical use,” Tomas Björklund, PhD, Lund University, Sweden, said in a press release.

Björklund is lead author of the study “A systematic capsid evolution approach performed in vivo for the design of AAV vectors with tailored properties and tropism,” which was published in the journal Proceedings of the National Academy of Sciences. 

The adeno-associated virus (AAV) is a common, naturally-occurring virus, which has been shown to work as an effective gene therapy delivery vehicle for genetic diseases, such as spinal muscular atrophy. In gene therapy, scientists deliver a working version of a faulty gene using a harmless AAV that was modified and inactivated in the lab. This way the virus functions only as a delivery vehicle and does not have the capacity to damage tissues and cause disease.

While AAVs have a natural ability to penetrate any cell of the body and infect as many cells as possible, their usefulness as a potential therapy requires the capacity to specifically deliver a working gene to a particular cell type, such as dopamine producing-nerve cells. Those are the ones hose responsible for releasing the neurotransmitter dopamine and that are gradually lost during Parkinson’s disease.

A team of Swedish researchers have developed a new method — called barcoded rational AAV vector evolution, or BRAVE — that combines powerful computational analysis with the latest gene and sequencing technology to produce AAVs that can specifically target neurons.

To make AAVs neuron specific, the team selected 131 proteins known to specifically interact with synapses (the junctions between two nerve cells that allow them to communicate).

They then divided the proteins into small sequences, called peptides, and created a large library where each peptide could be identified by a specific pool of genetic barcodes (a short sequence of DNA that is unique and easily identified).

The peptide is then displayed on the surface of the AAV capsid, allowing researchers to test the simultaneous delivery of many cell-specific AAVs in a single experiment.

The team then injected these AAVs into the forebrain of adult rats and observed that around 13% of the peptides successfully homed to the brain. Moreover, 4% of the peptides were transported effectively through axons (long neuronal projections that conduct electrical impulses) toward the nerve cell’s body.

Researchers then selected 23 of these unique AAV capsids and injected them into rats’ striatum, a brain region involved in voluntary movement control and affected in Parkinson’s disease. Twenty-one of the new AAV capsids had an improved transport capacity within nerve cells than in standard AAVs.

One particular capsid, called MNM008, showed a high affinity for rat dopaminergic neurons. Researchers then tested whether this viral vector also could target human dopaminergic neurons.

The team transplanted neurons generated from human embryonic stem cells into rats’ striatum. Six months later, they injected either MNM008 or a control AAV capsid and found that MNM008 was able to target these specific cells and be transported into dopaminergic neuronal cell bodies through axons.

“Thanks to this technology, we can study millions of new virus variants in cell culture and animal models simultaneously. From this, we can subsequently create a computer simulation that constructs the most suitable virus shell for the chosen application — in this case, the dopamine-producing nerve cells for the treatment of Parkinson’s disease,” Björklund said.

Overall, researchers believe the BRAVE method “opens up the design and development of synthetic AAV vectors expressing capsid structures with unique properties and broad potential for clinical applications and brain connectivity studies.”

The team has established a collaboration with a biotech company, Dyno Therapeutics, to use the BRAVE method in the design of new AAVs.

“Together with researchers at Harvard University, we have established a new biotechnology company in Boston, Dyno Therapeutics, to further develop the virus engineering technology, using artificial intelligence, for future treatments,” Björklund said.

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Dosing Begins in Phase 2 Trial of CNM-Au8, Potential Therapy for Dopaminergic Neurons

Parkinson's pilot trial

A pilot Phase 2 study evaluating CNM-Au8, an investigational Parkinson’s treatment aiming to protect nerve cells, has started dosing patients, the therapy’s developer, Clene Nanomedicine, announced.

The open-label REPAIR-PD (NCT03815916) clinical trial is enrolling up to 24 people, ages 30 to 80 and diagnosed within the past three years, at its one site at the University of Texas Southwestern Medical Center.

“We are excited to be advancing CNM-Au8 clinically into Parkinson’s patients starting with the REPAIR-PD Phase 2 study” Rob Etherington, president and CEO of Clene, said in a press release.

Parkinson’s disease is characterized by the death of dopaminergic neurons in two brain regions, the striatum and the substantia nigra. These nerve cells rely on large amounts of energy to function, which is provided by mitochondria, the cell’s powerhouses. Failure to provide the energy that cells need contributes to their death.

Oxidative stress, an imbalance between the production of harmful free radicals and the ability of cells to detoxify them, is another hallmark of Parkinson’s disease. These free radicals, or reactive oxygen species, are produced during certain metabolic reactions that involve mitochondria, and can damage cells.

CNM-Au8 is a suspension of nanocrystalline gold designed to increase the production of energy. Specifically, it works by increasing the speed of conversion between two molecules — nicotinamide adenine dinucleotide (NADH) to its oxidized form (NAD+) — resulting in greater production of energy in the form of ATP (adenosine triphosphate, an energy-carrying molecule of cells).

In addition, CNM-Au8 has antioxidant properties that may help to protect cells against oxidative stress.

Preclinical (in the lab) data showed that CNM-Au8 aided the survival of dopaminergic neurons, and helped prevent the loss of mitochondria.

In a rat model of Parkinson’s disease, treatment with CNM-Au8 improved the animal’s motor activity compared to control (untreated) mice. Of note, rats treated with CNM-Au8 in this test showed better results than did rats given carbidopa/levodopa, a standard Parkinson’s therapy.

“Our preclinical data with CNM-Au8 demonstrated improvements in neuronal bioenergetics, which may improve the survival of dopaminergic neurons in patients with PD [Parkinson’s disease], thereby helping slow the progression of this devastating disease,” Etherington said.

A Phase 1 clinical trial involving healthy volunteers (NCT02755870) found CNM-Au8 to be safe.

In the REPAIR-PD study, patients will first undergo a four-week screening period, after which they will drink two ounces of CNM-Au8 daily each morning for 12 weeks. Treatment will be followed by a four-week follow-up period.

The study’s primary outcome is to determine improvements in oxidative stress in the central nervous system (brain and spinal cord), assessed by the ratio of NAD+/NADH measured using magnetic resonance spectroscopy (MRS).

Additional (secondary) measures include assessing the effects of CNM-Au8 on energy production, and nerve cell metabolism.

“The objective of the REPAIR-PD Phase 2 study is to demonstrate improvements in brain bioenergetic metabolism in Parkinson’s disease patients treated with CNM-Au8. Participants will undergo 31phosphorous magnetic resonance spectroscopy (31P-MRS) imaging to show how treatment with CNM-Au8 results in improvements in brain metabolic and membrane biomarkers,” said Robert Glanzman, MD, chief medical officer of Clene.

Results from the REPAIR-PD trial are expected by mid-2020.

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Tiny Crystals May Help Scientists Trace Causes of Brain Inflammation in Parkinson’s, Study Reports

brain inflammation

Tiny and traceable man-made crystals, known as quantum dots, may be useful in carrying toxins to select cells in the brain, allowing researchers to better understand Parkinson’s neurodegenerative processes by being able to model and visualize them, researchers in Canada report.

Their study, “Quantum dot conjugated saporin activates microglia and induces selective substantia nigra degeneration,” was published in NeuroToxicology.

Microglia, primary immune cells of the brain and spinal cord, are known to contribute to the inflammation that underlies Parkinson’s neurodegeneration, which severely affects a brain area involved in motor control called the substantia nigra.

A key aspect of Parkinson’s research is to understand new and targeted ways of modulating microglia’s behavior, with a goal of influencing the survival or neurons or nerve cells. Such an ability would help to address the precise link between neurons and microglial cells.

Quantum dots, or nanoscale crystals, can be specifically taken up by microglia cells and may be useful as a direct way of targeting these cells. The nanoparticles also glow a particular color after being illuminated by ultraviolet light, allowing scientists to trace the molecules inside cells and study their cellular behavior.

Researchers at Carleton University investigated whether microglia within the substantia nigra of mice would take up quantum dots alone, and quantum dots carrying an immunotoxin called saporin. The latter works by inactivating ribosomes — cells’ protein builders — which compromises protein synthesis and leads to cell death. The scientists also studied how these nanoparticles affected microglia status (i.e., whether it is active or inactive).

Animals were given a four-minute infusion directly into their substantia nigra of either quantum dots alone, or of one of two doses of quantum dots conjugated with saporin. Within a week post-infusion, the mice’s balance and coordination were assessed.

Using imaging technology, researchers observed that quantum dots alone were selectively taken up by microglia and activated them. Microglia activation is a hallmark of inflammation in the context of neurodegenerative disorders. Despite their activated state, however, microglia had minimal effects on neurons and other neuronal support cells like astrocytes.

But in mice whose quantum dots were administered together with saporin, scientists observed a significant dose-dependent reduction in the number of nigral — meaning “of the substantia nigra” — neurons that produce dopamine, as well as impaired motor coordination six days after the infusion.

Quantum dots conjugated to saporin also increased the levels of a molecular mediator of inflammation, called WAVE2. This protein “is critical for the changes in activation state morphology of microglia,” the scientists wrote

The researchers believe that quantum dots carrying saporin could be a new and targeted way of modeling Parkinson’s-related inflammation, and evaluating new therapies aiming to treat it.

“[Quantum dots] might be a viable route for toxicant delivery and also has an added advantage of being fluorescently visible,” they wrote.

“Future work using this model should attempt to establish various degrees of neuronal loss. This model could then be used to test neuro-recovery or protective agents at differing stages of [the disease],” the researchers added.

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Targeting Dopaminergic Neurons in ‘Zombie’ State Might Slow Parkinson’s Progression, Study Says

senescence and neurons

Contrary to what is commonly thought, dopamine-producing nerve cells (neurons) that stop functioning in Parkinson’s disease may not die, but instead enter a state of senescence in which they cease to divide and cause damage to healthy neighboring cells, a study found.

In fact, one researcher noted these senescence cells are considered “zombie cells,” a spreading “undead.”  Future therapies that specifically stop senescence may help prevent the disease or slow its progression.

The study, “Loss of SATB1 Induces p21-Dependent Cellular Senescence in Post-mitotic Dopaminergic Neurons,” was published in the journal Cell Stem Cell.

A key hallmark of Parkinson’s disease (PD) is the progressive degeneration of dopaminergic neurons in the brain, resulting in its characteristic motor symptoms.

During the natural process of aging, the main risk factor for both the sporadic and genetic forms of PD, humans and other organisms accumulate senescent cells within their tissues. Cellular senescence refers to when cells cease to divide and grow, and can no longer regenerate tissues.

While cellular senescence is important in both embryonic development and wound healing, and plays a role in preventing the development of certain cancers (by arresting uncontrolled cell growth), it can become detrimental.

Senescent cells tend to emit chemicals into their environment that can damage surrounding cells. In addition to accumulating in healthy, older tissue, senescent cells can abnormally accumulate in disease states.

In fact, recent studies have reported increased markers of cellular senescence in the brains of Parkinson’s patients.

Special AT-rich sequence-binding protein 1 (SATB1) was recently identified as a risk factor for PD. Previous studies have shown that SATB1’s activity is reduced in the most affected brain regions of patients.

Researchers set out to investigate SATB1’s role in dopamine-producing neurons, whose activity is lower than usual in Parkinson’s disease.

The team differentiated human stem cells into dopaminergic neurons in a lab dish; in some of these neurons, they silenced the gene responsible for producing the SATB1 protein.

Genetic deletion of SATB1 induced senescence in dopaminergic nerve cells. In particular, researchers found that a lack of SATB1 led to the characteristic hallmarks of cellular senescence, such as increased oxidative protein damage, damaged mitochondria — a cell’s “powerhouse” or energy source — and enlarged nuclei.

Dopaminergic neurons lacking SATB1 also released certain molecules that caused inflammation and senescence in surrounding neurons.

Further analysis found that in healthy dopaminergic neurons, SATB1 directly binds to the regulatory region of the p21 gene and represses its expression. This gene produces a protein known to promote senescence. As such, in a healthy scenario, SATB1 prevents dopaminergic neurons from entering senescence.

Eliminating SATB1 from another type of neuron, called CTX neurons, did not induce senescence or affected p21 expression.

The researchers believe that “SATB1-dependent repression of [p21] transcription seems to be crucial for [dopaminergic] neuron function.”

These findings may explain why Parkinson’s patients experience a drop in dopamine levels before dopamine neurons actually die.

“They [dopaminergic neurons] loose the function of a neuron even though they are still there,” Markus Riessland, the study’s lead author, said in a press release. “People call these senescent cells zombie cells because they’re undead, basically, and because their dead-like appearance is spreading.”

Reducing the activity of SATB1 in dopaminergic neurons in mice also resulted in the same signs of senescence and high levels of p21 and a local immune reponse.

These senescent neurons “stop the cell cycle and they start secreting inflammatory factors that signal to the immune system, ‘Come here and eat me,’” Riessland explained. “This might really be a novel explanation for why you see certain markers of inflammation in Parkinson’s Disease.”

Importantly, researchers found that p21 is actively expressed in dopaminergic neurons of Parkinson’s patients with the sporadic form of the disease, making these cells more prone to enter into a state of senescence.

The team believes that SATB1 could be a promising target for novel therapies that target senescent cells, called senolytics, which have already been able to improve age-related manifestations in mice.

Importantly, therapeutic strategies that target SATB1 or p21 in Parkinson’s disease could be a “beneficial route to intervention.”

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Potential Therapy for Dopaminergic Neurons, CNM-Au8, Enter Phase 2 Trial in Parkinson’s Patients

CNM-Au8 studies

CNM-Au8, an investigational therapy by Clene Nanomedicine, improved the survival of dopaminergic neurons, helped prevent loss of mitochondria, and rescued motor function in a rat model of Parkinson’s disease, a study reports.

The effects of CNM-Au8 will now be assessed in an open-label (no placebo group) Phase 2 trial, called REPAIR-PD (NCT03815916). This pilot study is enrolling up to 24 patients who will undergo treatment at the University of Texas Southwestern Medical Center. More information can be found here.

Karen Ho, Clene’s head of translational medicine, presented the preclinical findings in the scientific poster, “Gold nanocatalysis as a novel therapeutic for neuroprotection in Parkinson’s disease,” during the recent 2019 Society for Neuroscience (SfN) Annual Meeting in Chicago.

Parkinson’s disease is characterized by the degeneration and death of a particular group of nerve cells — called dopaminergic neurons —  in two brain regions, the striatum and the substantia nigra.

To work as intended, these nerve cells require large amounts of energy, which is provided by mitochondria: small organelles within cells that work as their powerhouses. Failure to provide the energy cells need contributes to their death.

Another key player in Parkinson’s is oxidative stress, an imbalance between the production of harmful free radicals and the ability of cells to detoxify them. These free radicals, or reactive oxygen species, are produced during certain metabolic reactions in which mitochondria are involved, and damage cells.

CNM-Au8 is a suspension of nanocrystalline gold that acts to increase the speed of certain intracellular reactions. Specifically, CNM-Au8 is designed to increase the conversion of nicotinamide adenine dinucleotide (NADH) to its oxidized form (NAD+), resulting in greater production of ATP, a key energy-carrying molecule used by mitochondria. In addition, CNM-Au8 has antioxidant properties that may help to protect cells against oxidative stress.

In the preclinical study, researchers treated co-cultures of rat dopaminergic neurons and glial cells (cells that surround neurons and provide them with support) with CNM-Au8. This significantly increased the total intracellular levels of NAD+ compared to treatment with the control, a reaction that leads to greater release of ATP.

Exposing these co-cultures to two neurotoxins — substances that damage the nervous system and mimic what occurs in Parkinson’s — called MPP+ (1-methyl-4-phenylpyridinium) and 6-OHDA, led to damage and death of dopaminergic neurons. However, treatment with CMN-Au8 significantly increased these neurons’ survival and helped to preserve the network generated by nerve cells.

In cells exposed to 6-OHDA, use of CNM-Au8 led to fewer clumps of alpha-synuclein protein, an established hallmark of Parkinson’s disease.

CNM-Ai8 treatment also halted the accumulation of reactive oxygen species, and significantly lessened the loss of mitochondria induced by exposure to the neurotoxin MPP+.

“We are excited to share these latest Parkinson’s disease neuroprotection data regarding our lead nanocatalyst, CNM-Au8, with the neuroscience research community,” Rob Etherington, the president and CEO of Clene, said in a press release.

“Coupled with prior neuroprotection and remyelination data presented at major scientific congresses, this new Parkinson’s data demonstrate how improvements in bioenergetics with CNM-Au8 may preserve neuronal viability across multiple neurodegenerative disorders,” Etherington added.

CNM-Au8 was also tested in vivo (living organism) in a rat model of Parkinson’s disease. 6-OHDA was injected into the right side of the animals’ striatum. These rats were then given CNM-Au8 or a sham solution daily, delivered orally, beginning the next day (early treatment) or 14 days later (late treatment group).

Locomotor function in the rats was assessed using the vertical cylinder paw placement test after six weeks. Results showed that animals treated with CNM-Au8, either early or later, had improvements in motor activity compared to control (untreated) mice.

Early treatment with CNM-Au8 also reduced the number of apomorphine-induced rotations (circling, which signals problems) in rats with lesions at week six by 42% compared to control rats.

Notably, rats treated with CNM-Au8 in this test also showed better results compared to rats treated with carbidopa/levodopa, a standard Parkinson’s therapy.

“These data support our belief that treatment with CNM-Au8 may improve the survival of dopaminergic neurons in patients with PD, thereby helping slow the progression of this devastating disease. Disease modifying therapies remain a key, unmet treatment goal in Parkinson’s disease,” Etherington said.

CNM-Au8 was shown to be safe in a Phase 1 clinical trial involving healthy volunteers (NCT02755870).

In the REPAIR-PD study, participants will first undergo a four-week screening period, after which they will drink two ounces of CNM-Au8 daily each morning for 12 weeks. Treatment will be followed by a four-week follow-up period.

The study’s primary outcome is to determine improvements in oxidative stress in the central nervous system (brain and spinal cord), assessed by the ratio of NAD+/NADH measured using magnetic resonance spectroscopy (MRS).

Additional (secondary) measures include assessing the effects of CNM-Au8 on energy production and nerve cells’ metabolism. Results are expected by mid-2020.

“We are excited to be advancing CNM-Au8 into studies in Parkinson’s patients starting with the REPAIR-PD Phase 2 study,” said Robert Glanzman, Clene’s chief medical officer. “This study will advance our understanding how CNM-Au8 treatment affects central nervous system biomarkers related to bioenergetics, neuronal metabolism, and oxidative stress, as potential indicators of target engagement for CNM-Au8.”

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Specific Dopamine-producing Neurons Crucial to Adaptive Movement, Early Study Finds

motor skills and Parkinson's

Dopaminergic neurons — nerve cells gradually lost to Parkinson’s progression — that contain an enzyme called aldehyde dehydrogenase 1A1 are essential for acquiring the motor skills needed for proper movement in given situations, a mouse study reports.

The research, “Distinct connectivity and functionality of aldehyde dehydrogenase 1A1-positive nigrostriatal dopaminergic neurons in motor learning,” was published in Cell Reports. The work was developed by the Intramural Research Program of the National Institute on Aging (IRP-NIA).

Parkinson’s disease severely affects dopaminergic neurons, those that produce dopamine, a neurotransmitter (cell-signaling molecule) that relays information between nerve cells and between the brain and the rest of the body.  These neurons are found in two specific brain regions involved in motor control: the striatum and the substantia nigra.

Nerve cells may or not contain aldehyde dehydrogenase 1A1 (ALDH1A1), an enzyme that is involved in cellular detoxification. Parkinson’s seems to mostly damage ALDH1A1-positive dopaminergic neurons, suggesting the enzyme may be a key player in this neurodegenerative disorder.

Both ALDH1A1-positive and ALDH1A1-negative dopaminergic nerve cells contribute to voluntary motor behavior. But the degree to which ALDH1A1-positive neurons are crucial to acquiring motor skills remains to be understood.

Using a mouse model of Parkinson’s, scientists targeted  dopaminergic neurons positive for ALDH1A1, and produced a detailed connectivity map of these specific neuronal networks in the mouse brain.

ALDH1A1-positive neurons were found to be in constant communication with other brain structures there. Importantly, researchers found that those dopamine-producing neurons of the striatum and substantia nigra that received the greatest percentage of molecular information (input) were located in the caudate-putamen nuclei, a brain region involved in movement control.

Researchers then selectively removed ALDH1A1-positive neurons to mimic the degeneration pattern observed in late-stage Parkinson’s disease. The animals’ ability to show new motor skills — new ways of voluntary movement, like foot position for maintaining balance while walking on a moving surface — was assessed using the rotarod test. In this test, mice must learn to balance while walking on a constantly rotating rod much like a treadmill.

Mice without ALDH1A1-positive neurons displayed a distinctly poorer ability to learn new motor skills, and slower walking speeds compared to control animals.

“Compared with a modest reduction in high-speed walking, the ALDH1A1+ nDAN-ablated mice showed a more severe impairment in rotarod motor skill leaning,” the researchers wrote. “Unlike control animals … [these] mice essentially failed to improve their performance during the course of rotarod tests.” (nDANs are nigrostriatal dopaminergic neurons.)

These animals were then treated with dopamine replacement therapy, either levodopa or a dopamine receptor agonist, one hour before a new motor skills assessment. Dopamine replacement therapy is standard treatment for the motor symptoms associated with Parkinson’s.

Levodopa (L-DOPA) treatment allowed the animals without ALDH1A1-positive neurons to travel longer distances, and to walk more frequently at higher speeds during a session. But it failed to improve their ability to acquire new motor skills during repeated tests. Treatment with a dopamine receptor agonist was also ineffective.

“When the ALDH1A1+ nDANs were ablated after the mice had reached maximal performance, the ablation no longer affected the test results, supporting an essential function of ALDH1A1+ nDANs in the acquisition of skilled movements. These findings are in line with the theory that nigrostriatal dopamine serves as the key feedback cue for reinforcement learning,” the researchers wrote.

These results provide “a comprehensive whole-brain connectivity map,” they concluded, and reveal a key role of ALDH1A1-positive neurons in newly learned motor skills, suggesting that motor learning processes require these neurons to receive a multitude of information from other nerve cells and to supply dopamine with “dynamic precision.”

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