So, the glymphatic system is a very hot area of neuroscience right now, so we'll have Dr. Plagg come to the podium to tell us about his work looking at this in decompressive craniectomy. All right. Well, good afternoon. So my name is Ben Plagg. I'm a third-year medical student in the Medical Scientist Training Program at the University of Rochester, and the title of my talk this afternoon is going to be Decompressive Craniectomy Impairs Glymphatic Circulation Syndrome of the Trephined. I have no disclosures. So to start, I'll just go over a brief case. This is a case of a 48-year-old female who was involved in a traumatic sledding accident, and her head CT upon presentation demonstrated bilateral hematomas as well as diastasis of the coronal suture. She underwent bilateral craniotomies for decompression and went on to have a full neurologic recovery post-operatively. However, several months later, she developed a sudden onset of severe headaches as well as pseudobulbar palsy, and repeat head CT performed at that time demonstrated bilaterally sunken skin flaps. And so at this point, she underwent bilateral cranioplasty and, again, made a full neurologic recovery post-operatively, this time permanently. And so this is syndrome of the trephined, and so in addition to headaches as well as focal neurologic deficits, this syndrome has been characterized, or rather associated with a variety of neurologic signs and symptoms, including dizziness, mood changes, seizures, et cetera. And so the question is, why does this syndrome occur? There's been a number of hypotheses, maybe the most simple of which is just compression of the underlying cortex by an enfolded scalp. This is why this syndrome is sometimes referred to as syndrome of the sinking skin flap. Additionally, changes in cerebral blood flow have been hypothesized to play a role. But what our lab's most interested in is how alterations in cerebrospinal fluid and interstitial fluid hydrodynamics may contribute. And the reason we're interested in this, as was alluded to before, is our work in identifying and characterizing what we've termed the lymphatic system, which is a perivascular pathway for interstitial waste clearance within the brain. So just very briefly to kind of go over the anatomy and physiology of the system, what we've been able to demonstrate is that subarachnoid CSF penetrates into the brain parenchyma very specifically within periarterial spaces. And then interstitial fluid surrounding the cells of the brain drains from the brain parenchyma within perivenous spaces. And then aquaporin-4 water channels, which are densely localized to astrocytic end-foot processes that circumscribe the entirety of the brain's cerebrovasculature, help to facilitate movement of CSF from periarterial spaces into the interstitium, and then interstitial fluid into perivenous spaces. We've been able to show that convective bulk flow through this pathway is important for the clearance of extracellular solutes and metabolic waste products. And a breakdown of flow through this pathway has been implicated in diseases such as Alzheimer's and traumatic brain injury. And so this led us to ask, does decompressive craniectomy impair glymphatic CSF flow, and can this impairment in glymphatic flow produce syndrome of the trefined? So to address this question, we developed a very basic mouse model of syndrome of the trefined, where we removed a 2-millimeter diameter section of skull overlying the right parietal cortex, and then we also removed the underlying dura in these animals. And then in these craniectomy mice and a group of control mice, we introduced a fluorescently labeled protein tracer into the CSF via the cisterna magna, and we allowed it to circulate around and into the brain for a 30-minute period so as to assess glymphatic CSF influx kinetics. At 30 minutes, we perfusion fixed these animals, coronally sectioned their brains, and then used fluorescence microscopy to evaluate CSF penetration into the brains. And so what we can see here is that immediately, within 15 minutes of performing a decompressive craniectomy, there's a significant reduction in glymphatic CSF inflow in multiple different brain regions, ranging from the cerebral cortex to the hippocampus, subcortical structures, and white matter, both ipsilateral and contralateral to the decompressive craniectomy, suggesting that this process leads to a global or brain-wide suppression of CSF inflow. Because syndrome of the trefined is a more chronic condition that develops in a more delayed fashion, we were interested in whether or not there was persistent glymphatic failure that resulted from decompressive craniectomy. And similarly, or conversely, we wanted to know whether or not cranioplasty, putting the bone flap back in place, could help restore glymphatic function. So again, using our syndrome of the trefined model, there was a group of craniectomy mice which received a craniectomy on day zero. A subset of mice received a cranioplasty on day 14, and then we introduced our fluorescent protein tracer into the CSF on days 14, 28, and 56. And so what we can see is that by 14 days following decompressive craniectomy, there's a persistent, significant impairment of glymphatic CSF inflow, again, in multiple brain regions, both ipsilateral and contralateral. What we were surprised to find, however, is that by 28 days, there's a spontaneous recovery of glymphatic inflow, and this is irrespective of whether or not a cranioplasty had been performed. And so this is just a quantification of that data, and I'll just kind of move on for the sake of time for right now. So having demonstrated that there's a stagnation of glymphatic flow out to at least 14 days following decompressive craniectomy, we were interested in whether or not this had a consequence, specifically whether or not it could produce a neuroinflammatory response. So in our chronic craniectomy and cranioplasty models, we also performed immunohistochemistry for an astraglial marker of neuroinflammation, GFAP, and here we find that at 14 days following decompressive craniectomy, in the ipsilateral cortex, there's a significant elevation in GFAP expression. This persists out to 28 days, but what's interesting is that if cranioplasty is performed by 28 days, there's no significant elevation in GFAP expression. So what this suggests is that recovery in glymphatic function tends to precede any recovery from the neuroinflammatory response, and cranioplasty can accelerate this process. When we looked at a microglial marker of neuroinflammation, CD68, we found a very similar phenomenon. At 14 days, there was a significant elevation in CD68 expression, both ipsilateral and contralateral, in both ipsilateral and contralateral cortices. This persisted out to 28 days. If cranioplasty was performed, there was no longer a significant elevation of CD68. So finally, having demonstrated that decompressive craniectomy results in persistent glymphatic impairment, and this produces neuroinflammation, we were curious whether or not this would have a neurobehavioral consequence. Specifically, we looked at coordinated motor function using the Rotorod, and we looked at object memory looking at the novel object recognition test. And we performed these assessments at baseline before any surgery was performed, and then at 14, 21, and 28 days following craniectomy. What we find is that in terms of gross locomotor function, there's no significant differences in terms of mean speed or distance traveled amongst our groups. However, what was interesting is when we looked at coordinated motor function, not surprisingly, control mice are able to learn this task over a 28-day assessment period, allowing them to stay on the Rotorod for longer periods of time. Decompressive craniectomy mice are never able to learn this task, and therefore never improve in terms of their duration on the Rotorod. Cranioplasty mice start out just as bad as the craniectomy mice, but then by 28 days are able to stand the Rotorod at comparable durations to the controls. Similarly, when we looked at object memory, we can see that control mice are readily able to discern a novel object placed in their cage. Decompressive craniectomy mice aren't able to do this with any higher fidelity than random chance, so 50-50 chance. And our cranioplasty mice start out with an inability to identify the novel object, but by 28 days are able to identify the novel object at similar rates to our controls. So to summarize and conclude, what I've been able to demonstrate today is decompressive craniectomy leads to impaired glymphatic CSF influx. Glymphatic stagnation is associated with a neuroinflammatory response, as well as the development of motor and cognitive deficits. And finally, glymphatic recovery tends to precede any reduction in the neuroinflammatory response in resolution of motor and cognitive deficits, and cranioplasty accelerates this recovery. So here are just some acknowledgments. My PhD advisor, Dr. Niedergaard, Dr. Edvates, also, and everybody else who contributed to this project, the Medical Scientist Training Program, and my funding sources. Thank you. All right, wonderful. Thank you for that work.

glymphatic system

decompressive craniectomy

glymphatic flow

neuroinflammation

cognitive deficits