Intermittent fasting (IF) has been suggested to have neuroprotective effects through the activation of multiple signaling pathways. Rodents fasted intermittently exhibit enhanced hippocampal neurogenesis and long‐term potentiation (LTP) at hippocampal synapses compared with sedentary animals fed an ad libitum (AL) diet. However, the underlying mechanisms have not been studied. In this study, we evaluated the mechanistic gap in understanding IF‐induced neurogenesis.
We evaluated the impact of 3 months of IF (12, 16, and 24 hr of food deprivation on a daily basis) on hippocampal neurogenesis in C57BL/6NTac mice using immunoblot analysis.
Three‐month IF significantly increased activation of the Notch signaling pathway (Notch 1, NICD1, and HES5), neurotrophic factor BDNF, and downstream cellular transcription factor, cAMP response element‐binding protein (p‐CREB). The expression of postsynaptic marker, PSD95, and neuronal stem cell marker, Nestin, was also increased in the hippocampus in response to 3‐month IF.
These findings suggest that IF may increase hippocampal neurogenesis involving the Notch 1 pathway.
Dietary restriction (DR) is defined as a decrease in energy consumption without reducing nutritional value. This simple dietary intervention has been shown in a wide range of experimental animals to extend lifespan and decrease the incidence of several age‐related diseases. The definition of DR has been expanded from an alternative description of caloric restriction (CR) to also encompass a broader scope of interventions, including short‐term starvation, periodic fasting, fasting‐mimetic diets, and intermittent fasting (IF; Mattson & Arumugam, 2018). IF has been proven to be advantageous to various organ systems in the body and acts as a mild metabolic stressor. It has been postulated that IF is able to cause powerful changes in the metabolic pathways in the brain via an increase in stress resistance, and breakdown of ketogenic amino acids and fatty acids (Bruce‐Keller, Umberger, McFall, & Mattson, 1999; Kim et al., 2018). Experimental studies have also shown that IF is neuroprotective against acute brain injuries such as stroke, and neurodegenerative diseases (Arumugam et al., 2010; Halagappa et al., 2007; Manzanero et al., 2014). In addition, recent studies have also shown that IF can lead to an increase in neurogenesis levels in the hippocampus (Manzanero et al., 2014).
In the adult brain, the niches of neuronal stem cells (NSCs) are located specifically at the subventricular zone (SVZ) of the lateral ventricles, and in the subgranular zone (SGZ) of the hippocampus. The ability of NSCs to maintain cerebral neurogenesis is controlled by the tight regulation of balanced events commencing from stem cell maintenance, to stem cell division and proliferation, to its differentiation into mature neurons, and finally their survival and functional integration into the brain parenchyma (Lathia, Mattson, & Cheng, 2008; Lledo, Alonso, & Grubb, 2006). The process of adult neurogenesis is highly regulated and is adaptable to environmental, morphological, and physiological cues, whereby cerebral performance is suited to function at optimal levels for a given environment. Studies have demonstrated that the proliferation of neural stem cells can be modified through metabolic perturbations experienced during high temperatures (Matsuzaki et al., 2009), physical activity (Niwa et al., 2016), and a high‐fat diet (Kokoeva, Yin, & Flier, 2005). Experimental studies from our group have also shown that IF increases neurogenesis in the hippocampus as a form of neuroprotection following acute brain injury such as ischemic stroke. Moreover, we established that the number of BrdU‐labeled cells in the dentate gyrus of IF mice was elevated (Manzanero et al., 2014). To measure cell proliferation without the confound availability of an exogenous marker BrdU, we established increases in the number of Ki67‐labeled cells in the dentate gyrus of mice on the IF diet, indicating enhancement of cell proliferation in these mice (Manzanero et al., 2014). In addition to our findings, previous work similarly demonstrated that using the every other day (EOD) IF regimen also increased BrdU‐labeled cell number in the hippocampus (Lee, Duan, & Mattson, 2002).
However, the molecular process involved in IF‐induced neurogenesis is not well understood. The Notch signaling pathway that is intricately involved in the determination of cell fate during brain development and adult neurogenesis may be a possible molecular process involved in IF‐induced neurogenesis (Lathia et al., 2008). In this study, we investigated the expression levels of molecular and cellular components of the hippocampal region, focusing specifically on Notch activation and associated proteins that are known to promote hippocampal neurogenesis such as brain‐derived neurotrophic factor (BDNF) and cAMP response element‐binding protein (CREB).[…]
Imagine the brain could reboot, updating its damaged cells with new, improved units. That may sound like science fiction — but it’s a potential reality scientists are investigating right now. Ralitsa Petrova details the science behind neurogenesis and explains how we might harness it to reverse diseases like Alzheimer’s and Parkinson’s.
The evolutionary history of humans explains why physical activity is important for brain health
It is by now well established that exercise has positive effects on the brain, especially as we age.
Less clear has been why physical activity affects the brain in the first place.
Key events in the evolutionary history of humans may have forged the link between exercise and brain function.
Cognitively challenging exercise may benefit the brain more than physical activity that makes fewer cognitive demands.
In the 1990s researchers announced a series of discoveries that would upend a bedrock tenet of neuroscience. For decades the mature brain was understood to be incapable of growing new neurons. Once an individual reached adulthood, the thinking went, the brain began losing neurons rather than gaining them. But evidence was building that the adult brain could, in fact, generate new neurons. In one particularly striking experiment with mice, scientists found that simply running on a wheel led to the birth of new neurons in the hippocampus, a brain structure that is associated with memory. Since then, other studies have established that exercise also has positive effects on the brains of humans, especially as we age, and that it may even help reduce the risk of Alzheimer’s disease and other neurodegenerative conditions. But the research raised a key question: Why does exercise affect the brain at all?
Physical activity improves the function of many organ systems in the body, but the effects are usually linked to better athletic performance. For example, when you walk or run, your muscles demand more oxygen, and over time your cardiovascular system responds by increasing the size of the heart and building new blood vessels. The cardiovascular changes are primarily a response to the physical challenges of exercise, which can enhance endurance. But what challenge elicits a response from the brain?
Answering this question requires that we rethink our views of exercise. People often consider walking and running to be activities that the body is able to perform on autopilot. But research carried out over the past decade by us and others would indicate that this folk wisdom is wrong. Instead exercise seems to be as much a cognitive activity as a physical one. In fact, this link between physical activity and brain health may trace back millions of years to the origin of hallmark traits of humankind. If we can better understand why and how exercise engages the brain, perhaps we can leverage the relevant physiological pathways to design novel exercise routines that will boost people’s cognition as they age—work that we have begun to undertake.
FLEXING THE BRAIN
To explore why exercise benefits the brain, we need to first consider which aspects of brain structure and cognition seem most responsive to it. When researchers at the Salk Institute for Biological Studies in La Jolla, Calif., led by Fred Gage and Henriette Van Praag, showed in the 1990s that running increased the birth of new hippocampal neurons in mice, they noted that this process appeared to be tied to the production of a protein called brain-derived neurotrophic factor (BDNF). BDNF is produced throughout the body and in the brain, and it promotes both the growth and the survival of nascent neurons. The Salk group and others went on to demonstrate that exercise-induced neurogenesis is associated with improved performance on memory-related tasks in rodents. The results of these studies were striking because atrophy of the hippocampus is widely linked to memory difficulties during healthy human aging and occurs to a greater extent in individuals with neurodegenerative diseases such as Alzheimer’s. The findings in rodents provided an initial glimpse of how exercise could counter this decline.
Following up on this work in animals, researchers carried out a series of investigations that determined that in humans, just like in rodents, aerobic exercise leads to the production of BDNF and augments the structure—that is, the size and connectivity—of key areas of the brain, including the hippocampus. In a randomized trial conducted at the University of Illinois at Urbana-Champaign by Kirk Erickson and Arthur Kramer, 12 months of aerobic exercise led to an increase in BDNF levels, an increase in the size of the hippocampus and improvements in memory in older adults.
Other investigators have found associations between exercise and the hippocampus in a variety of observational studies. In our own study of more than 7,000 middle-aged to older adults in the U.K., published in 2019 in Brain Imaging and Behavior, we demonstrated that people who spent more time engaged in moderate to vigorous physical activity had larger hippocampal volumes. Although it is not yet possible to say whether these effects in humans are related to neurogenesis or other forms of brain plasticity, such as increasing connections among existing neurons, together the results clearly indicate that exercise can benefit the brain’s hippocampus and its cognitive functions.
Researchers have also documented clear links between aerobic exercise and benefits to other parts of the brain, including expansion of the prefrontal cortex, which sits just behind the forehead. Such augmentation of this region has been tied to sharper executive cognitive functions, which involve aspects of planning, decision-making and multitasking—abilities that, like memory, tend to decline with healthy aging and are further degraded in the presence of Alzheimer’s. Scientists suspect that increased connections between existing neurons, rather than the birth of new neurons, are responsible for the beneficial effects of exercise on the prefrontal cortex and other brain regions outside the hippocampus.
UPRIGHT AND ACTIVE
With mounting evidence that aerobic exercise can boost brain health, especially in older adults, the next step was to figure out exactly what cognitive challenges physical activity poses that trigger this adaptive response. We began to think that examining the evolutionary relation between the brain and the body might be a good place to start. Hominins (the group that includes modern humans and our close extinct relatives) split from the lineage leading to our closest living relatives, chimpanzees and bonobos, between six million and seven million years ago. In that time, hominins evolved a number of anatomical and behavioral adaptations that distinguish us from other primates. We think two of these evolutionary changes in particular bound exercise to brain function in ways that people can make use of today.
Researchers used to think that after adolescence, people were pretty well stuck with the brain cells they’d already formed. No so anymore. Discoveries in recent years have shown that neurogenesis—the formation of new neurons—can occur much later than this, well into adulthood. And now, a new study from the University of Illinois at Chicago finds that brain cells can form into one’s nineties, even if one has cognitive decline and Alzheimer’s disease (though at a much decelerated rate). The question is how the late-in-life growth of new neurons fits into what’s already known about degenerative diseases.
The study was published last week in the journal Cell Stem Cell.
The researchers looked at the postmortem brains of people aged 79-99, some of whom had had cognitive decline or Alzheimer’s disease. They targeted markers for two kinds of burgeoning cells—neuroblasts (stem cells that would one day give rise to neurons), and immature neurons—in the hippocampus, the brain area that’s most affected in Alzheimer’s disease.
People who had died without cognitive problems had proliferation of both kinds of cells in their brains. People with cognitive decline and Alzheimer’s also had evidence of the cells, but in much lower numbers.
Lazarov, neurogenesis study
COURTESY, ORLY LAZAROV, ET AL.
“We found that there was active neurogenesis in the hippocampus of older adults well into their 90s,” said study author Orly Lazarov in a statement. “The interesting thing is that we also saw some new neurons in the brains of people with Alzheimer’s disease and cognitive impairment.”
What was interesting was the finding that people who had scored higher on tests of cognition during their later lives had more neuroblasts in their hippocampi, compared to those who’d scored lower—and this was independent of the level of degeneration that was visible in the brain.
“In brains from people with no cognitive decline who scored well on tests of cognitive function, these people tended to have higher levels of new neural development at the time of their death, regardless of their level of pathology,” Lazarov said. “The mix of the effects of pathology and neurogenesis is complex and we don’t understand exactly how the two interconnect, but there is clearly a lot of variation from individual to individual.”
The finding is intriguing since it’s long been known that a person’s level of brain “gunk” (the plaques and tangles associated with Alzheimer’s disease) doesn’t always correlate with their cognitive and behavioral symptoms. So it’s possible that these new findings helps explain why this disconnect exists—perhaps the level of neurogenesis matters as much or more than the amount of plaques and tangles that develop. If that’s true, then the big question would be how to harness this for therapeutic purposes.
“The fact that we found that neural stem cells and new neurons are present in the hippocampus of older adults means that if we can find a way to enhance neurogenesis, through a small molecule, for example, we may be able to slow or prevent cognitive decline in older adults, especially when it starts, which is when interventions can be most effective,” said Lazarov.
More research will obviously be needed to understand all of this, but preventing cognitive decline and dementia is probably the way to go, especially since medications to treat Alzheimer’s after the fact have fallen flat in recent years. In the meantime, the study is encouraging on another level: Certain lifestyle habits—most notably exercise—have consistently been shown to boost neurogenesis. The findings suggest we’d do well to pick up exercise, and other brain-healthy habits, and engage in them for as much of our lives as we can, as regularly as we’re able.
New neurons are generated in the hippocampal dentate gyrus from early development through adulthood. Progenitor cells and immature granule cells in the subgranular zone are responsive to changes in their environment; and indeed, a large body of research indicates that neuronal interactions and the dentate gyrus milieu regulates granule cell proliferation, maturation, and integration. Following traumatic brain injury (TBI), these interactions are dramatically altered. In addition to cell losses from injury and neurotransmitter dysfunction, patients often show electroencephalographic evidence of cortical spreading depolarizations and seizure activity after TBI. Furthermore, treatment for TBI often involves interventions that alter hippocampal function such as sedative medications, neuromodulating agents, and anti-epileptic drugs. Here, we review hippocampal changes after TBI and how they impact the coordinated process of granule cell adult neurogenesis. We also discuss clinical TBI treatments that have the potential to alter neurogenesis. A thorough understanding of the impact that TBI has on neurogenesis will ultimately be needed to begin to design novel therapeutics to promote recovery.
Adult neurogenesis in the hippocampal dentate gyrus is widespread in mammals. Generation of dentate granule cells occurs late in embryonic development, continues after birth, and persists into old age in most mammals examined (Amrein et al., 2011; Amrein, 2015; Ngwenya et al., 2015). Studies in rodents indicate that adult generated granule cells play a role in hippocampal dependent learning (Nakashiba et al., 2012; Danielson et al., 2016; Johnston et al., 2016). Whether neurogenesis continues into old age in humans remains controversial (Danzer, 2018a), with studies finding evidence for (Eriksson et al., 1998; Spalding et al., 2013; Boldrini et al., 2018) and against ongoing neurogenesis (Sorrells et al., 2018). Yet there is general agreement that dentate neurogenesis occurs in childhood and continues throughout young adulthood in humans, and that newly-generated neurons are poised to contribute to hippocampal function. At a minimum, therefore, traumatic brain injuries (TBIs) occurring during adolescence have the potential to disrupt this important process.
The generation, maturation, and integration of new neurons is critical for hippocampal function. This tightly regulated process, however, is easily disrupted by pathological events, such as TBI. In this review, we discuss the coordinated process of adult neurogenesis in the hippocampal subgranular zone (SGZ) and the impact that TBI and TBI treatments have on this process. An understanding of the regulation and dysregulation of neurogenesis is important for determining whether and how therapeutic interventions targeted at adult neurogenesis are useful for TBI treatment.
Neurogenesis Is a Complex, Tightly-Regulated Process
Adult neurogenesis is characterized by multiple “control” points. The number of daughter cells produced by neural stem cells (NSC) located in the SGZ of the dentate gyrus can be modulated by the rate of cell proliferation and survival, while factors regulating fate specification control whether and how the new cells become neurons and integrate into the hippocampal circuitry (see recent review by Song et al., 2016). These control points can be regulated by signals released into the extracellular milieu by both neuronal and non-neuronal cells (Alenina and Klempin, 2015; Egeland et al., 2015), neurotrophic and transcription factors (Faigle and Song, 2013; Goncalves et al., 2016), neuroinflammatory mediators (Belarbi and Rosi, 2013), metabolic and hormonal changes (Cavallucci et al., 2016; Larson, 2018), and direct synaptic input from both glutamatergic and GABAergic neurons (Chancey et al., 2014; Alvarez et al., 2016; Song et al., 2016; Yeh et al., 2018). For additional information, the readers are referred to the excellent reviews cited for each mechanism, and the schematic in Figure 1. Critically, all of these factors can be disrupted by TBI, creating an environment in which immature granule cells and granule cell progenitors no longer receive the proper cues to guide their development.
Figure 1. Generation and integration of adult-born granule cells is a coordinated process that is impacted by TBI. At each stage of adult neurogenesis, the normal process (blue) has potential to be altered by TBI (orange). (1) Quiescent radial neural stem cells (NSCs) in the subgranular zone (SGZ) can be depleted by frequent activation early in life, such as by TBI-induced seizures, leading to deficiencies with age. (2) TBI and its effects, including spreading depolarizations and seizures, cause an increase in proliferation of progenitor cells. (3) Newly-generated neurons migrate from the SGZ to the granule cell layer (GCL), and after TBI abnormal hilar migration is apparent. (4) Parvalbumin interneurons and (5) mossy hilar neurons are susceptible to cell death after TBI. Reduction in their numbers results in decreased GABAergic and glutamatergic (respectively) input to the newly-generated neurons. Newly-generated neurons show additional signs of aberrant neurogenesis such as abnormal connectivity (6), hyperexcitability (7) and inappropriate integration and dendritic maturity (8) which can be caused by changes in the environmental milieu.
You can ask many different experts, and neuroplasticity will be explained in many different ways. The purpose of this website is not to get into technical jargon that overwhelms the stroke patient but rather to educate persons about stroke rehabilitation in simple laymen terms. In stroke recovery, neuroplasticity basically refers to the ability of the brain to rewire or reorganize itself after injury. Various studies over the past decade have shown that the adult brain can “rewire” itself when damaged. Studies have also shown that the adult brain can create new neurons, a phenomenon called neurogenesis. These new neurons require support from neighboring cells, blood supply, and connection with other neurons to survive. Certain requirements must be met during rehabilitation for neurogenesis and plasticity to actually change the brain. Rehabilitation involving neuroplasticity principles requires repetition of task and task specific practice to be effective. What this means for the stroke patient is that going to see your therapist for a one hour visit (or even a 3 hour visit) is not enough to lead to neuroplastic changes in the brain. Patients need to think of physical, occupational, and speech therapy as an adjunct to stroke recovery. It’s up to the patient to make the most of recovery by continuously using the injured parts of the body and mind outside of therapy sessions in everyday life.
A good comparison would be how one learns multiplication. A teacher doesn’t just show a multiplication table a couple of times to her students for the concept to be mastered. Instead, students have to practice over and over to learn and master multiplication. A child doesn’t learn how to walk overnight. It requires much practice. A baseball player doesn’t become elite just by playing a few games of baseball. You must take control of your stroke recovery process and be willing to invest a lot of time and energy if you want to see change especially with moderate to severe stroke. It’s also important to keep using a skill once you have mastered it – use it or lose it as you often hear in rehab.
Please note that plasticity doesn’t mean that one can practice every task over and over and accomplish them all. Stroke is much more complicated than that. Different parts of the brain control different body functions and the brain adapts better to some areas of damage more than others. Scientists have identified certain areas of the brain that yield neurogenesis but have not identified it in all areas of the brain. If you want to learn more about your specific stroke, ask your neurologist specifically what areas of your brain were affected. The neurologist will also be able to tell you what problems you can expect because of that damage (e.g. speech deficits, vision deficits, dizziness, difficulties with balance, etc.) You can further improve your rehabilitation by specifically targeting the weaknesses caused by your stroke.
In my opinion, neuroplasticity doesn’t necessarily change exercise and therapeutic activities done in stroke rehabilitation but rather emphasizes that more repetition and task specific practice is needed. Probably the most commonly used therapy that is based on neuroplasticity is constraint induced therapy. Constraint induced therapy involves limiting the movement of the non-affected or stronger arm and instead using the affected or weaker arm more frequently and intensely. There has been some positive research results with constraint induced therapy, however, it requires much effort and patience from the stroke patient. Some other treatments that may help with brain reorganization include interactive metronome, brain retraining software and websites, mirror box therapy, and robotic and gait devices that assist with movement repetition.
Research is still needed in the area of brain plasticity and stroke rehabilitation. Scientists have demonstrated that brain reorganization can occur, but only limited rehab treatments have been developed that address neuroplasticity. The stroke patient, however, armed with the knowledge that brain rewiring occurs with repetition, can improve their rehabilitation outcomes by application of this concept in their daily lives. Remember, therapy is an adjunct to recovery. You cannot go to therapy sessions and expect positive outcomes without applying what you have learned on a consistent daily basis.
NEURON NURSERY Roughly the same number of new nerve cells (dots) exist in the hippocampus of people in their 20s (three hippocampi shown, top row) as in people in their 70s (bottom). Blue marks the dentate gyrus, where new nerve cells are born. M. BOLDRINI/COLUMBIA UNIV.
Your brain might make new nerve cells well into old age.
The finding contradicts a study published in March, which suggested that neurogenesis in the hippocampus stops in childhood (SN Online: 3/8/18). But the new research fits with a larger pile of evidence showing that adult human brains can, to some extent, make new neurons. While those studies indicate that the process tapers off over time, the new study proposes almost no decline at all.
Understanding how healthy brains change over time is important for researchers untangling the ways that conditions like depression, stress and memory loss affect older brains.
When it comes to studying neurogenesis in humans, “the devil is in the details,” says Jonas Frisén, a neuroscientist at the Karolinska Institute in Stockholm who was not involved in the new research. Small differences in methodology — such as the way brains are preserved or how neurons are counted — can have a big impact on the results, which could explain the conflicting findings. The new paper “is the most rigorous study yet,” he says.
Researchers studied hippocampi from the autopsied brains of 17 men and 11 women ranging in age from 14 to 79. In contrast to past studies that have often relied on donations from patients without a detailed medical history, the researchers knew that none of the donors had a history of psychiatric illness or chronic illness. And none of the brains tested positive for drugs or alcohol, says Maura Boldrini, a psychiatrist at Columbia University. Boldrini and her colleagues also had access to whole hippocampi, rather than just a few slices, allowing the team to make more accurate estimates of the number of neurons, she says.
To look for signs of neurogenesis, the researchers hunted for specific proteins produced by neurons at particular stages of development. Proteins such as GFAP and SOX2, for example, are made in abundance by stem cells that eventually turn into neurons, while newborn neurons make more of proteins such as Ki-67. In all of the brains, the researchers found evidence of newborn neurons in the dentate gyrus, the part of the hippocampus where neurons are born.
Although the number of neural stem cells was a bit lower in people in their 70s compared with people in their 20s, the older brains still had thousands of these cells. The number of young neurons in intermediate to advanced stages of development was the same across people of all ages.
Still, the healthy older brains did show some signs of decline. Researchers found less evidence for the formation of new blood vessels and fewer protein markers that signal neuroplasticity, or the brain’s ability to make new connections between neurons. But it’s too soon to say what these findings mean for brain function, Boldrini says. Studies on autopsied brains can look at structure but not activity.
Not all neuroscientists are convinced by the findings. “We don’t think that what they are identifying as young neurons actually are,” says Arturo Alvarez-Buylla of the University of California, San Francisco, who coauthored the recent paper that found no signs of neurogenesis in adult brains. In his study, some of the cells his team initially flagged as young neurons turned out to be mature cells upon further investigation.
But others say the new findings are sound. “They use very sophisticated methodology,” Frisén says, and control for factors that Alvarez-Buylla’s study didn’t, such as the type of preservative used on the brains.
Deep in the hippocampus, are new neurons born throughout life? Just when scientists were about to reach some consensus that the answer was yes, two recent studies disagree. In the April 5 Cell Stem Cell, Maura Boldrini and colleagues at Columbia University, New York, report that adult neurogenesis not only exists, but remains steady into old age. The researchers counted newborn neurons in samples from people aged 14–79 years, and came up with similar numbers. In the March 7 Nature, researchers led by Arturo Alvarez-Buylla at the University of California, San Francisco, reported that while neural progenitors abounded in postmortem hippocampi from prenatal or early childhood brains, they fell off the map by age 7. What gives? In older people, some of the cells that expressed markers of budding neurons turned out to be glia, the authors claim.
Neural progenitor cells proliferate in human hippocampus throughout adulthood, says a new study.
It blames waning angiogenesis, not faulty neurogenesis, for lost neuroplasticity in old age.
In contrast, another paper claims brain neurogenesis fizzles during childhood.
It claims some cells bearing neural progenitor markers are actually glia.
Who is right? Researchers who spoke with Alzforum stood squarely behind Boldrini because she used stereology, a gold standard quantitation method, to estimate numbers of neural progenitors throughout the entire dentate gyrus of postmortem brain. Alvarez-Buylla’s team estimated cell numbers using only three to five slices from each postmortem sample. “It would be very difficult to rule out neurogenesis by this method,” said Orly Lazarov of the University of Illinois in Chicago, who pointed out that because the study relied on small numbers of individuals per age group, the data could be misleading.
Others, including Jonas Frisén of the Karolinska Institute in Stockholm, pointed to a long list of previous papers supporting the existence of neurogenesis in the adult human brain. “An analogy is that 10 people go into the woods to search for blueberries,” he wrote, “Nine come back with blueberries and one does not. Are there blueberries in that forest?”
Still, others acknowledged that some of those “blueberries” might have been glia. “To me, it boils down to a single question: are these proliferating cells truly neural progenitors, or not?” asked Costantino Iadecola of Weill Cornell Medical College in New York.
Numerous studies in rodents support the idea that neural progenitors in the mammalian brain continue to trickle out fresh neurons into adulthood, though factors such as aging and disease dampen the flow (Altman and Das, 1965; Sep 2001 news; Feb 2002 news; Kempermann et al., 2003; Mar 2010 news). Tracking neurogenesis in humans has been trickier, but a seminal study two decades ago set the stage: Five terminal cancer patients received an injection of bromodeoxyuridine (BrdU), a dye that incorporates into DNA during cell division. Postmortem analyses revealed evidence of dividing neurons in the dentate gyri of all of the patients (Eriksson et al., 1998). Since then, carbon-14 tracing and immunohistochemistry studies have supported the idea that new neurons arise in the adult human brain (Knoth et al., 2010; Jun 2013 news; Feb 2014 news).
Boldrini and colleagues set out to determine if age affects adult neurogenesis. They acquired hippocampal samples from 11 women and 17 men, aged 14–79, who were cognitively normal, had suffered no brain trauma, had had no microvascular pathology in the brain, and had clean toxicology reports at the time of death. The researchers collected 50-micron thick sections every 2 mm along the entirety of the hippocampus, and used both immunofluorescence and immunocytochemistry to label various cell-surface markers associated with five different stages of neural development (see image below). Finally, they estimated cell numbers throughout the dentate gyrus using stereology, whereby a computer algorithm calculates total cell numbers in a region by combining data from multiple sections. They reported the number of neural progenitors in the anterior, mid-, and posterior dentate gyrus.
The earliest neural progenitors known, called quiescent radial-like type I neural progenitors (QNPs), express GFAP, a marker shared with astrocytes; nestin, an intermediate filament protein that marks neural stem cells; and the transcription factor Sox2, which is required for the maintenance of multipotent stem cells. The researchers found that numbers of these cells decreased with age in the anterior-mid dentate gyrus. This is in keeping with the prevailing view that people are born with a finite number of these QNPs, Boldrini said.
QNPs give rise to type II intermediate neural progenitors. INPs are proliferating cells that express Ki67, a marker of actively dividing cells. Neuroblasts, or type III INPs, also proliferate, but lose expression of GFAP and Sox2. Based on expression of Ki67, nestin, and Sox2, the researchers determined that numbers of type II and III INPs remained steady, on the order of thousands of cells, in all three regions of the dentate gyrus throughout life. These neural progenitors were found in the subgranular zone (SGZ), which is proposed to be the predominant neurogenic niche in the region, as well as the granule cell layer (GCL, see image below). The findings pointed to a stable supply of neural progenitors in the dentate gyrus throughout adult life.
The researchers next asked whether those progenitors would fulfill their destiny and give rise to immature neurons and, ultimately, bona fide granule neurons. On the way to becoming fully fledged neurons, type III INPs start to express doublecortin (DCX), a microtubule-associated protein involved in neural migration. They also produce polysialylated neural cell adhesion molecule (PSA-NCAM), a glycoprotein they need for plasticity. Together, DCX and PSA-NCAM mark young neurons, which continue to express both proteins until they differentiate into mature neurons, whereupon they suppress DCX. The researchers found that the tissue donors had similar numbers of cells co-expressing DCX and PSA-NCAM, regardless of their age, suggesting neurogenesis continued unabated throughout life. Numbers of NeuN+ mature neurons also held steady, indicating that neuronal loss in the dentate gyrus is not a characteristic of healthy aging, either.
The researchers calculated that each dentate gyrus had between 10,000 and 15,000 young neurons (i.e., type III INPs and immature neurons). While the functional significance of these cell numbers is unclear, Boldrini speculated that this ongoing level of neurogenesis influences neural circuitry and cognition. For this reason, boosting neurogenesis could be a therapeutic strategy for neurodegenerative disease, she said.
However, while older adults appear to generate as many new neurons as younger people, those new cells may be less plastic, judging by a decline in PSA-NCAM+/DCX– cells in the anterior dentate gyrus. Curiously, using endothelial markers and stereology to measure the numbers, length, bifurcations, and total volume of capillaries, the scientists also found an age-dependent decline in angiogenesis in the same regions. The researchers proposed that a decline in angiogenesis may trigger loss of neuroplasticity without necessarily affecting neurogenesis, for example by starving new neurons of essential growth factors or nutrients.
Others were not convinced, noting that reliance on a single marker—PSA-NCAM—made the plasticity results no more than an interesting correlation. Still, Lazarov and Iadecola said the connection between age-related decline in angiogenesis and neuroplasticity was plausible. Iadecola was surprised that loss of angiogenesis did not appear to affect neurogenesis, but he noted that the donors had no obvious vascular pathology in their brains. Perhaps in people with more severe vascular problems, neurogenesis would be affected, he said.
In Grown-Up Brain, Nary a Newborn Neuron
In the Nature paper, first author Shawn Sorrells and colleagues used many of the same markers—Sox2, GFAP, DCX, and PSA-NCAM—to assess neurogenesis in postmortem samples across the lifespan. This included 11 samples from prenatal donors, the youngest of whom was only at 14 weeks gestation. They also analyzed seven samples from infants who died during their first year of life, one from a 7-year-old, one from a 13-year-old, and 17 samples from adults up to 77 years of age at the time of death. The samples came from multiple sources, and were not limited to healthy donors, or all postmortem. They included hippocampal tissue from surgical resection in 22 people with epilepsy, who ranged from three months to 64 years old.
For the postmortem samples, the researchers used three to five coronal sections to assess cell numbers. Rather than using stereology to estimate the total number of cells in the dentate gyrus, the researchers counted cells in individual sections. Three researchers independently counted each section while blinded to the age of the donor. They identified key structural landmarks, most notably the cell-dense GCL, to infer the relative locations of the cells.
In prenatal samples, the scientists found abundant proliferating Ki67+ cells that expressed the progenitor markers Sox1 and Sox2. Numbers of these cells plummeted during the first year of life, and were near zero in samples from people 7 or older. Notably, these proliferating cells never coalesced beneath the GCL to form a distinctive layer in the SGZ, a structural niche that supports neurogenesis in rodent models. The researchers confirmed the absence of this layer by electron microscopy on a subset of their samples, ranging in age from 22 gestational weeks to 48 years of age.
DCX+/PSA-NCAM+ cells, representing intermediate neural progenitors and immature neurons, clustered throughout the GCL at birth to a density of about 1,600 cells per mm2. In prenatal and infant samples, these cells had a smooth, elongated morphology characteristic of young neurons. By 13 years of age, sections only contained around two young neurons per mm2, or roughly one or two cells per section. Likewise, the investigators found no evidence of young neurons in samples from epilepsy patients older than 11. As for adults, none of the surgical or postmortem samples contained DCX+/PSA-NCAM+ cells, however the researchers did find cells that expressed PSA-NCAM without DCX. Unlike the elongated young neurons in infant samples, these cells had a more mature neuronal morphology with distinct axons and dendrites, and expressed NeuN, suggesting they were highly plastic neurons. The researchers also identified DCX+ cells in some older childhood and adult samples, but these cells co-expressed glial markers, and under the gaze of electron microscopy, had glial morphology.
The researchers also looked for evidence of neurogenesis in rhesus macaques. By staining with similar neuronal markers, they found that unlike in the human brain, proliferating neural progenitors did gather in the SGZ before birth. However, the number of these young neurons decreased eightfold between birth and 1.5 years of age, and were sparse in 7-year-old animals. Similarly, labeling dividing cells with BrdU revealed a steep drop-off in dividing neurons between 1.5 and 7 years of age.
The researchers concluded that neurogenesis is robust only in the earliest stages of development, and that DCX+ cells in late childhood and adult samples were actually glia. In an email to Alzforum, Sorrells and Alvarez-Buylla speculated that the cells identified as young neurons in the Boldrini study were also likely non-neuronal. “Identifying new neurons is technically challenging—in our own recent study we made similar observations to what Boldrini et al. report, but after extensive additional analysis of the shape and appearance of the cells in question, including electron microscopy and gene expression, we determined that these cells were not in fact young neurons or neural progenitors but different types of cells altogether,” they wrote.
However, Boldrini asserted that in her study, the cells stained for both DCX and PSA-NCAM did not co-localize with cells that appeared to be glia based on the pattern of Nissl staining, and were present in the thousands. Boldrini added that the immature neurons took on a pyramidal shape, characteristic of neurons, not glia.
Sorrells and Alvarez-Buylla further drew attention to the lack of a defined layer of proliferating cells in the SGZ in their study, adding that Boldrini’s samples also appeared to lack a distinct layer of cells there. In rodents, neural progenitors gather and proliferate in the SGZ. On this issue, Boldrini thinks that perhaps in humans neurogenesis occurs in a more scattered fashion. She said that for this reason, taking stock of cells throughout the entire dentate gyrus is crucial to capture these sparse cells.—Jessica Shugart