September 2016

Discovering How a Neurological ‘Pit Crew’ Keeps the Brain on Track

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Imagine taking neuroscientists to a NASCAR race. While most spectators keep their eyes on the speeding cars, you might catch a few scientists in the crowd instead watching the activities of the pit crews at work on the sidelines, helping drivers to refuel and repair their cars to keep racing at top form. Michael Robinson, PhD, would be one of them; metaphorically, research in his lab at The Children’s Hospital of Philadelphia has followed the actions of a neurological “pit crew” with research on the functions of astrocytes, the most abundant type of cell in the brain. His team is making early-stage foundational discoveries that could profoundly influence how scientists think about how the brain works, and eventually, how they treat brain injuries and neurological diseases.

“This work has changed a lot for us,” said Dr. Robinson, a professor of Pediatrics and Systems Pharmacology and Translational Therapeutics at CHOP and the Perelman School of Medicine at the University of Pennsylvania. “I’ve been here for 28 years. This whole story is something that people are really interested in. It is very gratifying to see people so excited.”

The excitement stems in large part from the fact that scientists are only recently awakening to the importance of astrocytes. Although neurons, like race-car drivers, usually get more attention for being the site of the action in brain functions, astrocytes play a vital supporting role in refueling and repair. Astrocytes respond to the energy demands of active neurons by relaying signals to blood vessels in the brain to send more oxygen-rich blood flow. Astrocytes also supply neurons with numerous vital chemicals and clear others to facilitate crisp signaling and prevent neuronal death. While it is widely recognized now that astrocytes have these essential “pit crew” functions in support of neurons, many unknowns remain about their specific activities and how they achieve them.

New insights and even more questions are emerging now thanks to the overturning of a misconception about astrocytes within the last decade. These bushy-shaped cells have multiple spiky protrusions (called processes) reaching out into the synapses between neurons. Until just a few years ago, scientists believed that mitochondria, the structures that are the primary source of energy within cells, were only located in the astrocytes’ cell body because the processes seemed too narrow to fit them.

“Now, I think most people agree that there are mitochondria throughout these processes in vivo and in vitro,” Dr. Robinson said. “Several different labs have now seen it. That essentially changes the equation.”

Finding out what mitochondria do in the spiky outer reaches of astrocytes became a key question. One obvious answer is that they help capture more of the energy from glucose in the form of ATP, which is the classic core function of mitochondria in all cells.

By investigating the movements of mitochondria within astrocytes, a team led by Joshua Jackson, PhD, then a postdoctoral fellow in Dr. Robinson’s lab, with Dr. Robinson and graduate student John O’Donnell, found another activity for these energizing organelles. Their results, published in the Journal of Neuroscience in 2014, implied that mitochondria support astrocytes’ ability to act like a cleanup crew to remove a hazard from the race track. Specifically, mitochondria appear to be recruited to positions within the astrocyte that are both near synapses and adjacent to transporter proteins that remove glutamate (the most common neurotransmitter in the brain); this removal is essential because glutamate is toxic to neurons with prolonged exposure.

Next, O’Donnell turned his attention to how mechanisms involving astrocytes are involved in the brain’s response after stroke. In the aftermath of a stroke, astrocytes undergo certain morphological changes — but, unlike a lot of neurons, they generally survive. This has made them a compelling potential target for future drug therapies. The team’s paper published this summer in the Journal of Neuroscience details the results of in vitro experiments to understand the activities of astrocytes and their mitochondria after a simulated stroke. O’Donnell, who defends his doctoral dissertation in Pharmacology in September and will next join the lab of D. Kacy Cullen, PhD, at Penn, was the paper’s first author.

One major finding was that mitochondria in the astrocytes’ spiky processes were broken down. This was a large effect, and one that was previously unknown, involving a cellular compartment that is vital for clearing glutamate from synapses. More surprisingly, they found that blocking uptake of glutamate into astrocytes prevented that loss of mitochondria. But the researchers still know too little to interpret these changes; they do not even know if the loss of mitochondria from astrocytes is good or bad for the brain’s recovery from stroke.

“It seems that during stroke, pathologic activation of glutamate transport is driving the loss of mitochondria,” Dr. Robinson said. “I have no idea what it means.”

In addition, the researchers learned more about how mitochondria are involved in calcium signaling within astrocytes, both under normal conditions and after the simulated stroke. Calcium signaling is a key mechanism by which astrocytes call for increased blood flow to the brain (a mechanism called neurovascular coupling). The team found that, under normal conditions, mitochondria are associated with two kinds of calcium signals, both of which are highly structured. A continuous cloud of calcium ions surrounds mitochondria, representing a low-level, constant hum of conversation in areas of the astrocyte process where mitochondria are located. And, intermittently, there is a so-called extramitochondrial spike, or an instantaneous calcium signal that extends between two mitochondria within the cell — a kind of quick text message between distant neighbors.

After a simulated stroke, O’Donnell and colleagues found that those two normally well-ordered calcium signaling patterns go haywire. The calcium clouds that once surrounded mitochondria shrink as many mitochondria break down, and the remaining mitochondria get smaller. Also, the extramitochondrial spikes are no longer contained between mitochondria, but propagate on and on. The exact impact of these altered signals is unclear, but the researchers feel confident that it has effects on neurovascular coupling.

Dr. Jackson, now a staff scientist at CHOP, is embarking on in vivo imaging to understand the mechanisms by which astrocytes control blood flow. And that is just the first of many questions that are unfolding to understand an area of basic science that is largely unknown territory.

“It’s great to have a strong mitochondrial research group at CHOP; it makes it easier for us to learn about the field and get feedback on our findings,” Dr. Robinson added. “The great thing about this job is that I’m always learning new fields. I just can’t believe that we’re sitting on this right now. The next steps are pretty obvious; it will be fun to see how it works.”

Because this research is still in early stages of basic science discovery, drug approaches for brain injury and neurological disease based on these findings are likely a long way away. Still, the potential for future therapies is something many investigators are attuned to as they learn more about astrocytes.

“Astrocytes are the most abundant cell type in the brain, so it may make sense to target these cells that are so well positioned, being completely integrated with synapses and the vasculature, and that aren’t that badly harmed during this injury,” O’Donnell said.

In other words, equipping the pit crew with the resources it needs might be a good way to help a race car driver get back on track after a crash.

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