Cerebral Hemispheres 2


How many glial cells are there in the brain?

One of the fundamental principles of scientific thinking is skepticism. A good scientist refuses to accept anything blindly, instead scrutinizing every purported statement of fact to make sure the evidence backs it up.

Because this mindset is so pervasive in scientific disciplines, it’s difficult to understand how unsubstantiated claims can be accepted as fact in science. But this does happen occasionally. Some unproven assertions have even found their way into the vaunted territory of common knowledge— a designation that means something is so well-established as true that you don’t even need a source to back it up anymore. For the last several decades, this has been the status of claims that there are many more glial cells than neurons in the brain.

Neuroscientists have been interested in finding accurate estimates of the number of neurons and glia in the brain for at least a century and a half. While figuring these numbers out is a prodigious feat no matter how you cut it, determining glial cell counts has been particularly challenging due to the small size of glia and the difficulty in telling them apart from other small cells. Still, cell counting methods have improved drastically over the last several decades, and there’s reason to believe that we finally have some valid estimates of both neurons and glial cells.

Methods for counting brain cells

Despite technical limitations like poor microscope resolution and undeveloped approaches to staining cells, early neuroscientists sometimes still managed to arrive at credible counts of neurons in the brain. Helen Bradford Thompson, for example, published an estimate in 1899 of the number of neurons in the cerebral cortex (about 9 billion) that matches up well with current estimates of about 10-20 billion.

Early neuroscientists like Helen Bradford Thompson arrived at neuronal numbers by actually counting neurons. In fact, this approach is still used today, just in a more refined manner. But the overall idea is the same: count the number of cells in various samples of brain tissue and extrapolate the numbers obtained to a larger brain region, or the whole brain.

A more recently developed method of cell counting uses some additional steps to make the process a bit easier and more precise. It involves taking a sample of brain tissue and homogenizing it—destroying the cell membranes, leaving the nuclei intact, and creating a soup-like mixture of liquefied brain. The nuclei can be stained with a fluorescent dye, antibodies can be used to differentiate between neural and non-neuronal cells, and then the nuclei can be counted.

This process is called isotropic fractionation. Isotropy is uniformity, and refers to the mixture formed after homogenization of the brain tissue. And fractionation indicates that cells are counted in a fraction of the whole tissue, and then the results are used to infer numbers in the rest of the brain region.

Gial cell estimates

Isotropic fractionation is a relatively new method. Before it was developed, finding accurate cell numbers in the brain was more painstaking and susceptible to errors. And, as mentioned above, glial cells were especially problematic.

This difficulty in counting glia was represented in some of the uncertainty researchers expressed about the number of glial cells in the brain before the 1980s. Although it was widely believed that the tiny glia outnumbered neurons, there was not a lot of hard evidence to prove this was the case. So it wasn’t uncommon to find scientists using qualifiers like "perhaps" when they made statements about glial cell counts. A common estimate at this time was that there were "perhaps" ten times as many glial cells as neurons.

But there were some who made more definitive statements. Holger Hyden, for example—a reputable biochemist and neuroscientist—stated more decisively in the 1960s that there are ten times as many glial cells as neurons. It’s probably the case that Hyden based his proclamations on specific regions of the brainstem he was studying where glia really do outnumber neurons significantly. But the extrapolation to the entire brain was nevertheless speculative, even though it was stated conclusively.

As can happen in scientific writing, researchers found Hyden’s declarations and others like them and cited them when writing journal articles or textbooks. Over time, this happened enough that the statements, which should never have been definitive, became common knowledge.

By the 1980s, even the most reputable sources in neuroscience were asserting that there are at least ten times as many glia as neurons in the brain. For example, in the 1985 edition of the famous neuroscience textbook, Principles of Neural Science (sometimes called the “bible of neuroscience”), it’s stated that glia outnumber neurons anywhere from 10 to 50 times. Because the text also estimated the number of neurons in the brain to be around 1 trillion (now considered a huge overestimate), the number of glia are implied to be somewhere between 10 and 50 trillion.

But again these numbers were all speculative, and they didn’t match up with the data researchers who were actually counting cells had obtained. For example, a respectable appraisal of the number of neurons and glia in the brain was published in 1986, and it suggested there are about 70-80 billion neurons and 40-50 billion glial cells. The largest number of glial cells reported in a primary research report was 130 billion in 1968.

These seemingly more accurate estimates, however, were largely ignored. It wasn’t until the late 2000s, when researchers began publishing data using isotropic fractionation, that the field took note of the discrepancies.

Debunking the myth

The groundbreaking paper in this respect was published by the Brazilian neuroscientist Suzana Herculano-Houzel and her colleagues in 2009. They used the isotropic fractionation method to count neurons and glia in the brain, and ended up with estimates of 86 billion neurons and 85 billion non-neuronal cells (which included glia and other cells, like endothelial cells). This suggested that there were actually fewer glial cells than neurons, which agreed with some of the data obtained earlier.

There was a bit of resistance to accepting these numbers at first, as some argued that isotropic fractionation had not yet been validated by comparing its results with those obtained through more well-known cell-counting methods. These validation studies came with time, however, and subsequent studies supported the numbers of Herculano-Houzel’s group. Today, most researchers have accepted the data obtained with isotropic fractionation, and the preponderance of the evidence supports the idea that the ratio of glia to neurons is about 1:1.

Of course, this doesn’t diminish the importance of glia. Historically, they have not been given due credit for the integral roles they play in the brain. That seems to be changing in recent years, however, as we learn more about the functions of glia. And as we get a more accurate view of all that glia do, we also seem to be letting go of inaccurate estimates of their numbers.

Perhaps more than anything, the history of glial cell counting teaches us that we should be skeptical of any sources that make claims that aren’t directly supported by primary research. Just because an authoritative source says something definitively, it doesn’t necessarily mean that it’s true or even that the research backs it up. It’s important, especially in this age of extreme information availability, that we be highly critical of the information we consume.

Reference (in addition to linked text above):

von Bartheld CS, Bahney J, Herculano-Houzel S. The search for true numbers of neurons and glial cells in the human brain: A review of 150 years of cell counting. J Comp Neurol. 2016 Dec 15;524(18):3865-3895. doi: 10.1002/cne.24040. Epub 2016 Jun 16.


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