Equal numbers of neuronal and nonneuronal cells make the human

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The Journal of Comparative Neurology 513:532–541 (2009)
Equal Numbers of Neuronal and Nonneuronal Cells Make
the Human Brain an Isometrically Scaled-Up Primate
Brain
FREDERICO A.C. AZEVEDO,1 LUDMILA R.B. CARVALHO,1 LEA T. GRINBERG,2,3 JOSÉ MARCELO FARFEL,2
RENATA E.L. FERRETTI,2 RENATA E.P. LEITE,2 WILSON JACOB FILHO,2 ROBERTO LENT,1
1
AND SUZANA HERCULANO-HOUZEL *
1
Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Cidade Universitária, Ilha do Fundão 21941-590
Rio de Janeiro, Brazil
2
Grupo de Estudos em Envelhecimento Cerebral da Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil
3
Instituto Israelita de Ensino e Pesquisa Albert Einstein
ABSTRACT
The human brain is often considered to be the most cognitively capable among mammalian brains and to be much
larger than expected for a mammal of our body size. Although the number of neurons is generally assumed to be a
determinant of computational power, and despite the widespread quotes that the human brain contains 100 billion
neurons and ten times more glial cells, the absolute number
of neurons and glial cells in the human brain remains unknown. Here we determine these numbers by using the
isotropic fractionator and compare them with the expected
values for a human-sized primate. We find that the adult
male human brain contains on average 86.1 ⴞ 8.1 billion
NeuN-positive cells (“neurons”) and 84.6 ⴞ 9.8 billion NeuN-
negative (“nonneuronal”) cells. With only 19% of all neurons
located in the cerebral cortex, greater cortical size (representing 82% of total brain mass) in humans compared with
other primates does not reflect an increased relative number of cortical neurons. The ratios between glial cells and
neurons in the human brain structures are similar to those
found in other primates, and their numbers of cells match
those expected for a primate of human proportions. These
findings challenge the common view that humans stand out
from other primates in their brain composition and indicate
that, with regard to numbers of neuronal and nonneuronal
cells, the human brain is an isometrically scaled-up primate
brain. J. Comp. Neurol. 513:532–541, 2009.
© 2009 Wiley-Liss, Inc.
Indexing terms: human; brain size; neuron numbers; glia/neuron ratio; evolution; comparative
neuroanatomy
It is repeatedly stated in the literature and in neuroscience
textbooks that the human species is an unusually encephalized primate species, whose brain, five to seven times larger
than expected for a mammal of its body size (Jerison, 1973;
Marino, 1998), contains 100 billion neurons and about ten
times more glial cells (Kandel et al., 2000; Ullian et al., 2001;
Doetsch, 2003; Nishiyama et al., 2005; Noctor et al., 2007). The
supposedly unusual scaling of the human brain, however,
derives from comparisons across orders (Jerison, 1973) and,
even when restricted to primates, regards only the brain– body
relationship (Marino, 1998) rather than addressing how its
cellular composition compares with that expected from other
primates.
Moreover, to our knowledge, the widespread numbers on
the cellular composition of the human brain have never been
supported by experimental studies. The high anisotropy and
large size of the human brain hinder stereological determination of cell numbers and their distribution in the brain as a
whole. Estimates of the cellular composition of the human
© 2009 Wiley-Liss, Inc.
brain are available only for some structures, such as the
cerebral cortex (von Economo and Koskinas, 1925; Shariff,
1953; Pakkenberg, 1966; Pakkenberg and Gundersen, 1997;
Pelvig et al., 2008), cerebellum (Lange, 1975; Andersen et al.,
1992), and some subcortical nuclei (Pakkenberg and Gundersen, 1988). Such studies have estimated the number of
Grant sponsor: FAPERJ (to S.H.-H., R.L.); Grant sponsor: CNPq (to S.H.-H.,
R.L.); Grant sponsor: Pronex (to R.L.); Grant sponsor: CAPES (to R.E.P.L.);
Grant sponsor: IIEP-Albert Einstein (to L.T.G.); Grant sponsor: Alexander
von Humboldt Foundation (to L.T.G.).
The last two authors contributed equally to this work.
*Correspondence to: Suzana Herculano-Houzel, Instituto de Ciências
Biomédicas, Universidade Federal do Rio de Janeiro, Av. Brigadeiro
Trompowski s/n, Ilha do Fundão 21941-590 Rio de Janeiro-RJ, Brasil.
E-mail: [email protected]
Received 6 June 2008; Revised 15 September 2008; Accepted 16 December 2008
DOI 10.1002/cne.21974
Published online in Wiley InterScience (www.interscience.wiley.com).
The Journal of Comparative Neurology
THE HUMAN BRAIN AS A SCALED-UP PRIMATE BRAIN
cells in the human cerebral cortex as 3, 7, 14, 19 –23, or 21–26
billion neurons and, very recently, 28 –39 billion glial cells
(Pelvig et al., 2008), and the number of cells in the human
cerebellum has been estimated as 70 or 101 billion neurons
(Lange, 1975; Andersen et al., 1992) and fewer than 4 billion
glial cells (Andersen et al., 1992). From such studies, the total
number of neurons in the human brain might be inferred to fall
anywhere between about 75 and 125 billion plus an undetermined number of neurons in the brainstem, diencephalon, and
basal ganglia that may or may not be comparatively small.
Additionally, no evidence is found to support the common
quote of ten times more glial cells than neurons in the human
brain. The glia:neuron ratio in subcortical nuclei can be as high
as 17:1 in the thalamus (Pakkenberg and Gundersen, 1988),
but, given the relatively small combined number of glial cells
reported for the cerebral (Pelvig et al., 2008) and cerebellar
(Andersen et al., 1992) cortices, the only possible explanation
for the quote of ten times more glial than neuronal cells in the
entire human brain would be the presence of nearly one trillion
glial cells in the remaining structures.
We have recently determined the cellular scaling rules that
apply to the brain of a number of rodent and primate species
and found that, whereas the rodent brain increases in mass
faster than it gains neurons (defined as NeuN-positive cells)
across species, suggesting that the average neuronal cell size
increases in larger rodent brains (Herculano-Houzel et al.,
2006), the primate brain increases in mass linearly with increases in its number of neurons across species, suggesting
that the average neuronal cell size does not increase significantly with brain size (Herculano-Houzel et al., 2007). The
power laws relating body mass, brain mass, and number of
neurons for rodent and primate species allowed us to predict
that, if built according to the cellular scaling rules that apply to
rodents, a brain of 100 billion neurons should weigh over 45 kg
and belong to a body of 109 tons. In contrast, if built according to the scaling rules that apply to primates, this brain of 100
billion neurons should weigh 1.45 kg and belong to a body of
73 kg, values that approach those observed in humans, suggesting that the human brain is indeed constructed according
to the same rules that apply to other primates.
We thus set out to determine the total cellular composition
of the human brain with the aid of the same method (the
isotropic fractionator; Herculano-Houzel and Lent, 2005) and
relying on the same criterion of NeuN labeling to identify
“neurons” and “nonneuronal cells” in order to evaluate how its
composition compares with the expected composition of a
primate brain of its size. While supporting several independent stereological estimates, our results challenge the values
so often cited in the literature and suggest that, with regard to
brain cellular composition, humans are just scaled-up, large
primates.
MATERIALS AND METHODS
Human material
All brains were obtained from the Brain Bank of the Brazilian
Aging Brain Study Group (Grinberg et al., 2007), located at the
University of São Paulo Medical School (FMUSP). The project
was approved by the Ethics Committee for Research Projects
Analysis (CAPPesq) of FMUSP, Research Protocol number
285/04. Informed consent for removal of the brains was pro-
533
vided by next of kin, who also responded to the Clinical
Dementia Rating Scale (CDR) semistructured interview and to
the Informant Questionnaire on Cognitive Decline in the
Elderly—Retrospective Version (IQCODE; Jorm and Jacomb,
1989; Morris, 1993). Four brains from 50-, 51-, 54-, and 71year-old males, deceased from nonneurological causes and
without cognitive impairment (CDR ⴝ 0, IQCODE ⴝ 3.0), were
analyzed. The brain of the 71-year-old male was included in
the analysis because it contained a similar number of cells
and an even slightly higher number of neurons than the other
brains. The corpses remained at 4°C until the brains were
removed from the cranium less than 24 hours after death and
fixed immediately.
Fixation and dissection
Brains were fixed by perfusion with 4 liters of 2%
phosphate-buffered paraformaldehyde through the basilar artery and the internal carotids, followed by immersion for 36
hours in the same fixative. Fixation for less than 48 hours was
critical to allow for antibody recognition of NeuN, while still
being enough to guarantee that the nuclei remained intact
throughout the homogenization procedure. The meninges and
major blood vessels were removed, and the brains were split
sagitally into two hemispheres. After dissecting each cerebellar hemisphere by cutting the cerebellar peduncles at the
surface of the brainstem, each hemisphere was cut into 1-cmthick coronal sections, and the cerebral cortex was separated
from the remaining regions (basal ganglia, diencephalon,
mesencephalon, and pons, named collectively “rest of brain,”
or RoB) by cutting through the white matter along the surface
of the striatum in each section. In three of the four brains
analyzed, one of the hemispheres of the cerebral cortex had
the gray matter dissected away from the underlying white
matter by careful shaving of the gray matter around the gyri
with a scalpel until the white matter was exposed. The medulla was excluded because of inconsistency in the inferior
section level among cases during the autopsy procedure.
After fixation, the three main regions of interest (cerebellum,
cerebral cortex, and RoB) were stored in phosphate-buffered
saline (PBS; pH 7.4) at 4°C and subjected individually to the
isotropic fractionator method (Herculano-Houzel and Lent,
2005). Each structure was cut into smaller pieces that could
be homogenized in a tissue grinder and counted in 1 day, and
partial results were added together. Determining the total
number of cells in each brain typically required about 4 – 6
weeks.
Isotropic fractionator
The isotropic fractionator method has been described elsewhere (Herculano-Houzel and Lent, 2005). Briefly, it consists
of a chemomechanical dissociation of fixed biological tissue
in a saline detergent solution (1% Triton X-100, 40 mM sodium
citrate) using 40 –200-ml glass tissue grinders, followed by
intense agitation of the suspension containing all nuclei in the
original structure, in order to achieve isotropy. After adding
the fluorescent DNA marker 4ⴕ-6-diamino-2-phenylindole dihydrochloride (DAPI) to the suspension, the density of nuclei
is quantified by use of a hemocytometer under a fluorescence
microscope (Fig. 1A,C). The total number of nuclei is calculated by multiplying the density of nuclei by the total suspension volume and heretofore is referred to as “total number of
cells” in each structure.
The Journal of Comparative Neurology
534
F.A.C. AZEVEDO ET AL.
Figure 1.
Aspect of the nuclei in the hemocytometer. A,B: Typical low-magnification fluorescent micrographs of the same field of cerebellar cell nuclei
in suspension stained with DAPI (A) and for NeuN immunoreactivity (B). The arrowheads indicate nuclei that are NeuN negative and therefore
identified as nonneuronal nuclei. All other nuclei are NeuN positive and therefore identified as neuronal. Note that nuclei are intact and well
scattered. C,D: High-magnification confocal image of NeuN-negative (arrowheads; arrow, nonneuronal nucleus undergoing cell division) and
NeuN-labeled cerebellar cell nuclei. The clear, debris-free preparation of free cell nuclei makes the anti-NeuN immunoreactivity easy to
distinguish from the virtually nonnexistent background. Scale bars ⴝ 40 ␮m in B (applies to A,B); 20 ␮m in D (applies to C,D).
Immunocytochemistry
Neuronal nuclei from an aliquot of the suspension were
selectively immunolabeled overnight, at room temperature,
with mouse monoclonal anti-NeuN antibody (Chemicon, Temecula, CA; MAB377B clone A60 against murine NeuN;
Mullen et al., 1992) at a dilution of 1:200 in PBS. This
antibody is increasingly used in the literature as a neuronal
marker in qualitative (see, e.g., Eriksson et al., 1998; Cossette et al., 2007; Fajardo et al., 2008) as well as quantitative
(Gittins and Harrison, 2004; Dawodu and Thom, 2005) stud-
ies of the human brain. Anti-NeuN clone A60 labels no glial
cells and recognizes all neuronal cells of most, though not
all, subtypes in a variety of vertebrate species, including
humans (Mullen et al., 1992; Wolf et al., 1996; Sarnat et al.,
1998; Lyck et al., 2008). Neuronal subtypes in the central
nervous system known to present no labeling for NeuN
include Purkinje cells, mitral cells of the olfactory bulb,
inferior olivary and dentate nucleus neurons (Mullen et al.,
1992), neurons in the substantia nigra pars reticulata of the
gerbil (but not of the rat; Kumar and Buckmaster, 2007) and
The Journal of Comparative Neurology
THE HUMAN BRAIN AS A SCALED-UP PRIMATE BRAIN
possibly others, as yet unidentified. Here we identify and
count as “neurons” all NeuN-stained nuclei and count as
“nonneuronal cells” all nuclei that lack NeuN labeling. Although the numbers of neurons in the cerebellum and RoB
(which includes the inferior olive) are thus necessarily underestimated, the number of nonstained neurons included
therefore in the population designated “nonneuronal” is
likely to be very small and actually insignificant compared
with the total numbers of cells in these structures
(Andersen et al., 1992). A thorough analysis of adjacent
sections of human cerebral cortex stained with cresyl violet
or NeuN has shown that both methods give correlated
estimates of neuronal density, indicated that NeuN is particularly useful for distinguishing small neurons from glia
and confirmed the value of NeuN as a tool for quantitative
neuronal morphometric studies in human brain tissue (Gittins and Harrison, 2004). Additionally, because we identified
labeled nuclei by visual inspection under the microscope
and not by automated methods, we could confirm that all
NeuN-labeled nuclei in each sample were indeed of neuronal morphology and that all nuclei of a particular labeled
morphology were labeled in the sample.
After the nuclei were washed in PBS, they were incubated
for 2 hours at room temperature with AlexaFluor 555 antimouse IgG secondary antibody (Molecular Probes, Eugene,
OR), at a dilution of 1:200 in PBS in the presence of 10%
normal goat serum. The neuronal fraction in each sample
was estimated by counting NeuN-labeled nuclei in at least
500 DAPI-stained nuclei. NeuN staining is smooth, covers
the entire nuclear area, and is crisp and easily identifiable
from the very low background (Fig. 1B,D). The total number
of neurons in each structure was calculated by multiplying
the fraction of nuclei expressing NeuN by the total number
of nuclei. The number of nonneuronal nuclei was obtained
by subtraction. Photomicrographs for documentation were
taken using a Zeiss Axioplan fluorescence microscope or,
for high magnification, a Zeiss LSM 510 Multiphoton microscope and were acquired digitally in AxioVision or LSM
Image Browser software (all from Carl Zeiss MicroImaging),
respectively. For illustrations, contrast and brightness of
the micrographs were adjusted in Corel Draw X3.
Data analysis
All statistical analyses and regressions were performed in
StatView software (SAS, Cary, NC). All data reported are
mean ⴞ SD.
RESULTS
We find that the male human brain, aged ⬃50 years (n ⴝ 3)
or 70 years (n ⴝ 1) and weighing 1,508.91 ⴞ 299.14 g, contains
on average 170.68 ⴞ 13.86 billion cells. Among these, 85.08 ⴞ
6.92 billion cells are located in the cerebellum, 77.18 ⴞ 7.72
billion cells are in the cerebral cortex (including both gray and
white matter), and 8.42 ⴞ 1.50 billion cells are found in the
remaining regions (RoB; Fig. 2). Because no significant differences were found in mass or in neuronal, nonneuronal, and
total cell numbers between right and left hemispheres (t-test,
P values typically well above 0.1), all numbers given refer to
the combined hemispheres.
Overall, the nonneuronal/neuronal ratio in the whole human
brain is close to 1 (Fig. 2), insofar as half of the cells in the
535
human brain, or 86.06 ⴞ 8.12 billion, are neurons (range 78.82–
95.40 billion neurons). The fractional distribution of neurons in
the human brain does not correspond to the fractional distribution of mass among brain structures (Figs. 2, 3). Although
82% of brain mass consists of cerebral cortex (including subcortical white matter) and 42% consists of cerebral cortical
gray matter alone, the 16.34 ⴞ 2.17 billion neurons found in
this structure represent only 19% of all brain neurons. In
contrast, the cerebellum, which represents only 10% of total
brain mass, contains 69.03 ⴞ 6.65 billion neurons, or 80% of
all neurons in the human brain. Fewer than 1% of all brain
neurons are located in the RoB, comprising basal ganglia,
diencephalon, and brainstem (Figs. 2, 3), although this percentage is necessarily underestimated as a result of the
known lack of NeuN staining in at least some structures in the
brainstem (see Materials and Methods).
The other half of all human brain cells, or 84.61 ⴞ 9.83
billion, are nonneuronal cells, yielding a nonneuronal/
neuronal ratio of 0.99 for the human brain as a whole (Fig.
2). Nonneuronal cells outnumber neuronal cells in the RoB,
with a nonneuronal/neuronal ratio of 11.35 (Fig. 2). In the
gray matter of the cerebral cortex, the nonneuronal/
neuronal ratio is 1.48. In contrast, in the cerebellum, this
ratio is only 0.23 (Fig. 2).
In contrast to the distribution of neurons, the fractional
distribution of nonneuronal cells in the brain resembles more
closely the fractional distribution of mass in the structures
(Fig. 3). The cerebral cortex, including the subcortical white
matter, holds 72% (or 60.84 ⴞ 7.02 billion) of all nonneuronal
cells in the brain, whereas the cerebellum has 19% (or 16.04 ⴞ
2.17 billion) and the RoB holds 9% (or 7.73 ⴞ 1.45 billion) of all
the nonneuronal cells in the brain.
The separate analysis of white matter (WM) and gray
matter (GM) of the cerebral cortex shows that the former
contains 69.6%, or 19.88 ⴞ 2.83 billion, of the nonneuronal
cells of the cerebral cortex of one hemisphere (Fig. 4). The
ratio between nonneuronal and neuronal cells is 1.48 ⴞ 0.42
for the GM (Fig. 4) and 3.76 ⴞ 0.55 for the combined GM and
WM (Fig. 2).
The numbers of neuronal and nonneuronal cells found in
the human brain fall very close to the values expected for a
primate brain of human dimensions built according to the
linear, isometric cellular scaling rules found to apply to
primate brains (Herculano-Houzel et al., 2007; Fig. 5a– c).
According to these rules, a primate brain weighing 1,508 g
would be expected to have 94 billion neurons, with 22
billion neurons in the cerebral cortex, 78 billion neurons in
the cerebellum, and 0.6 billion neurons in the RoB (Table 1).
The observed numbers of neurons are actually slightly
smaller than expected in the human cortex and cerebellum
and larger in the RoB (Table 1). The percentage deviations
in the observed values from the numbers of neuronal and
nonneuronal cells expected in each structure of the human
brain given its mass fall within the same range as the values
found for each of the species from which the cellular scaling
rules were originally obtained (Herculano-Houzel et al.,
2007; Fig. 6). The neuronal and nonneuronal cell densities
found in the human brain also fall in the range observed in
nonhuman primates in that study (Herculano-Houzel et al.,
2007; Fig. 5d,e).
The Journal of Comparative Neurology
536
F.A.C. AZEVEDO ET AL.
Figure 2.
Absolute mass, numbers of neurons, and numbers of nonneuronal cells in the entire adult human brain. Values are mean ⴞ SD and refer to the
two hemispheres together. B, billion.
Figure 3.
Distribution of mass, numbers of neurons, and numbers of nonneuronal cells in the adult human brain. Each bar represents the percentage mass
or percentage number of cells located in the cerebral cortex (Cx, black), cerebellum (Cb, gray), and remaining areas of the brain (RoB, white).
Values are mean ⴞ SD.
DISCUSSION
It is often stated in the literature that glial cells outnumber
neurons in the human nervous system by a factor of 10. Our
finding that the human brain has an approximately 1:1 ratio of
nonneuronal:neuronal cells implies a necessarily smaller glia/
neuron ratio, if endothelial and other mesenchymal cells were
removed from the nonneuronal pool. Although this ratio of
approximately 1:1 for the whole brain is important counterevi-
The Journal of Comparative Neurology
THE HUMAN BRAIN AS A SCALED-UP PRIMATE BRAIN
537
Figure 4.
Absolute mass, numbers of neurons, and numbers of nonneuronal cells in the cortical gray and white matter. Values are mean ⴞ SD and refer
to the right hemisphere (RH) only (n ⴝ 3).
dence to the common overestimation of numbers of glial cells
in the brain, it conceals the fact that specific structures of the
human brain can have maximal glia/neuron ratios (if all nonneuronal cells were glial cells) as small as 0.23, such as the
cerebellum, and as large as 11.35, such as the RoB. The
maximal glia/neuron ratio of 1.48 that we observed in the total
GM of the cerebral cortex is close to the ratio of 1.65 observed
by Sherwood et al. (2006) in layer II/III of human prefrontal area
9L and similar to the ratios between 1.2 and 1.6 that Pelvig et
al. (2008) encountered in the whole human neocortical GM.
These similarities corroborate our findings.
According to the common view in the literature, the glia:
neuron ratio increases with brain size (Reichenbach, 1989),
leading to a predominance of glial cells in large brains (Nedergaard et al., 2003) that would be compatible with a 10:1 ratio
in humans. We have shown that, although the average nonneuronal cell size is relatively invariant across brain structures
and species, an increasing predominance of glial cells with
brain size is indeed found in rodents (Herculano-Houzel et al.,
2006), in which average neuronal size increases together with
neuronal number, but not in the primates examined so far
(Herculano-Houzel et al., 2007), in which average neuronal
size, as with average nonneuronal cell size, is estimated to
remain relatively stable as numbers of neurons increase. All
primates we have analyzed until now (Herculano-Houzel et al.,
2007) exhibit ratios of nonneuronal/neuronal cells that are
similar to the approximately 1:1 ratio found in the human brain
as a whole.
The glia/neuron ratio has been considered to be of great
relevance because of the multiple functional relationships recently demonstrated between these cell types (Nedergaard et
al., 2003; Shaham, 2005) and has been hypothesized to reflect
neuronal activity (Reichenbach, 1989). Alternatively, the glia:
neuron ratio might simply follow the ratio between average
neuronal and average glial cell mass. We have proposed
(Herculano-Houzel et al., 2006) that this occurs as the nearly
all-neuronal parenchyma is invaded in early postnatal development by glial progenitors that divide until the newly formed
glial cells, of a relatively constant average size, reach confluence (Zhang and Miller, 1996). In this scenario, a neuronal
parenchyma of a given volume built of a large number of small
neurons is invaded by progenitors that will give rise to the
same number of glial cells as a parenchyma of the same
volume built of a smaller number of larger neurons, but the
latter, with larger neurons, will have a much larger glia:neuron
ratio than the former. It is noteworthy that the low glia:neuron
ratio in the cerebellum is due not to a conspicuous lack of glial
cells but rather to a very large number of very small neurons,
because its nonneuronal cell density is even somewhat larger
than that in other structures. The relative constancy of glial
cell densities observed across rodents and primates, humans
included, may therefore be more physiologically meaningful in
The Journal of Comparative Neurology
538
F.A.C. AZEVEDO ET AL.
Figure 5.
The human brain conforms to the cellular scaling rules that apply to primates. a– c: Mass of the cerebral cortex (Cx, solid circles), cerebellum
(Cb, open circles), RoB (triangles), and whole brain (crosses) of six primate species, tree shrews, and humans (h) as a function of body mass
(a), number of neurons (b), and number of nonneuronal cells (c). The power functions plotted refer to nonhuman primates only (Herculano-Houzel
et al., 2007), and all have exponents close to 1.0. Note that the data points for the human brain fall very close to the plotted functions. d,e:
Neuronal (d) and nonneuronal (e) densities in the different brain structures of six primate species, tree shrews, and humans (h) plotted against
structure mass.
The Journal of Comparative Neurology
THE HUMAN BRAIN AS A SCALED-UP PRIMATE BRAIN
TABLE 1. Observed and Expected Cellular Composition of the Human Brain
According to the Cellular Scaling Rules for Primate Brains1
For a primate of 75 kg
Total brain mass (g)
Total number of brain cells
Total number of brain neurons
Total number of brain nonneurons
For a primate brain of 1,508 g
Total number of neurons
Total number of nonneurons
For a primate cortex of 1,233 g
Total number of neurons
Total number of nonneurons
For a primate cerebellum of 154 g
Total number of neurons
Total number of nonneurons
For a primate RoB of 118 g
Total number of neurons
Total number of nonneurons
1
Expected
Observed
Difference
1,362
170.97
78.08
94.28
1,508
170.68
86.06
84.61
ⴙ10.7%
–0.2%
ⴙ10.2%
–10.2%
93.82
113.17
86.06
84.61
–8.3%
–25.2%
22.36
99.02
16.34
60.84
–26.9%
–38.6%
77.94
11.26
69.03
16.04
–11.4%
ⴙ42.4%
0.62
7.17
0.69
7.73
ⴙ11.3%
ⴙ7.8%
Results are given in billions.
Figure 6.
Deviation from the expected cellular composition of the brain structures in six nonhuman primate species and man. Each box depicts the
median deviation (expressed as percentage of the expected value)
and the 25th and 75th percentiles of the observed numbers of neuronal and nonneuronal cells in the cerebral cortex, cerebellum, and
remaining areas from the values expected according to the cellular
scaling rules derived from the six first species in the graph. The bars
indicate the 10th and 90th percentiles for each species, and the dots
indicate the upper and lower extreme deviations. Note that the human
brain deviates in its cellular composition from that expected for a
primate of its brain size as much as the other species from which the
cellular scaling rules for primate brains were derived.
terms of their role in the metabolic maintenance and functional support of brain tissue than their numeric ratio to neurons.
Our estimate of an average total of 86 billion neurons in the
human brain is compatible with previous stereological determinations for individual structures such as the cerebral cortex
(von Economo and Koskinas, 1925; Pakkenberg and Gundersen, 1997; Pelvig et al., 2008) and the cerebellum (Lange,
1975; Andersen et al., 1992). Exact numbers are probably
highly variable among humans, particularly given the variation
of over 50% in the number of cortical neurons among individuals of the same sex described recently in the literature (Pelvig
et al., 2008). Although the numbers of neurons that we report
539
for the RoB (which includes NeuN-negative neurons in the
inferior olive and possibly other structures as well) are necessarily underestimated, the number of NeuN-negative neurons
included in the “nonneuronal” population of the RoB is likely
to be negligible compared with the 85 billion neurons found in
the ensemble of cerebral cortex and cerebellum (because the
RoB is found to contain a total of only about 8 of the 170 billion
cells in the human brain), or to the 690 million neurons in the
RoB. Moreover, even in the improbable scenario in which all
cells in the RoB, composed of massive fiber tracts, were
NeuN-negative neurons, they would still amount to only 5% of
all brain cells.
More important than the exact number of neurons in the
human brain, however, are the implications of how this number compares with that expected for a primate brain of human
proportions. We have shown before that a brain with about
100 billion neurons built according to the cellular rules that
apply to scaling rodent brains would weigh 45 kg (HerculanoHouzel et al., 2007), well above the largest known whale brain.
Humans of 70 kg of body mass built according to these rules
would be expected to have a brain of only 145 g, instead of
1,500 g. That is, humans do indeed have a brain that is about
ten times larger and holds seven times more neurons than
predicted for a nonprimate mammal of its body size.
Remarkably, however, here we find for the first time that
the human brain conforms to the scaling rules observed for
a given group of mammals: six other primate species
(Herculano-Houzel et al., 2007), the only ones so far whose
total brain cellular composition is known. We show that
humans, despite the large relative size of the cerebral cortex, hold only 19% of all brain neurons in this structure, as
do other primates and rodents of different brain sizes, and
demonstrate that the mass and cellular composition of the
human brain deviate from the values expected for a primate
of 75 kg by only 10%. Given that the cellular scaling rules
that apply to primate brains are linear, the conformity of the
human brain to these rules strongly indicates that the human brain is a linearly scaled-up primate brain in its cellular
composition. The isometric scaling of the human brain cellular composition relative to other primates is in line with
other observations that have established that the cerebellum (Frahm et al., 1982) and frontal cortex (Semendeferi et
al., 2002) of the human brain have the same relative size as
in great apes, even though the distribution of mass within
the WM and individual cortical areas in humans may differ
from that in other primates (Semendeferi et al., 2001; Rilling
and Seligman, 2002; Schoenemann et al., 2005).
Our notion that the human brain is a linearly scaled-up
primate brain in its cellular composition is in clear opposition to the traditional view that the human brain is 7.0 times
larger than expected for a mammal and 3.4 times larger
than expected for an anthropoid primate of its body mass
(Marino, 1998). However, such large encephalization is
found only when body-brain allometric rules that apply to
nonprimates are used, as stated above, or when great apes
are included in the calculation of expected brain size for a
primate of a given body size. Our finding thus suggests that
the rules that apply to scaling brains are more conserved
than those that apply to scaling the body and raises the
intriguing possibility that, rather than humans having a
larger brain than expected, it is the great apes such as
The Journal of Comparative Neurology
540
F.A.C. AZEVEDO ET AL.
orangutans and, more notably, gorillas that have bodies
that are much larger than expected for primates of their
brain size. Indeed, the inclusion of great apes (Marino,
1998) in the primate species (Herculano-Houzel et al., 2007)
that we compare to humans would increase the body size
expected of our species, with a brain of 1,509 g, from 77 kg
to 216 kg, and decrease the expected brain size for a body
of 70 kg from 1,247 g to 557 g. One piece of evidence in
support of the possibility that gorillas and orangutans,
rather that humans, are outlier species in terms of body size
is that, whereas in most primate species, humans included,
the brain represents about 2% of total body mass (Marino,
1998), the brains of gorillas and orangutans, at about 500 g
(Semendeferi and Damasio, 2000), represent at most 1% of
a body of 50 kg, and only 0.5% or less in typical male
gorillas of 100 kg or more. The adaptive value of an enlarged body size in these species can be appreciated from
the status of social dominance that comes with the large
investment of time and energy necessary to develop large
bodies in alpha-male gorillas and orangutans (Leigh, 1995).
We are currently investigating whether the brains of gorillas and orangutans also conform to the cellular scaling
rules found to apply to other primates, including humans.
Such conformity would substantiate the intriguing possibility that, rather than humans having too large a brain for their
bodies, gorillas have too large a body for their brains,
although both species have brains built according to the
same rules that apply to other primates. Body size (Jerison,
1973), after all, may not be a relevant parameter when it
comes to cognition (Roth and Dicke, 2005). In light of the
recent finding that absolute brain size is the parameter that
best correlates with cognitive abilities (Deaner et al., 2007),
our cognitive advantage over other primates might be simply a consequence of having the largest brain, built with an
isometrically enlarged number of neurons compared with
other smaller-brained primates, regardless of body size.
ACKNOWLEDGMENTS
We thank all of our colleagues who helped with tissue
collection and homogenization.
LITERATURE CITED
Andersen BB, Korbo L, Pakkenberg B. 1992. A quantitative study of the
human cerebellum with unbiased stereological techniques. J Comp
Neurol 326:549 –560.
Cossette M, Lévesque D, Parent A. 2005. Neurochemical characterization
of dopaminergic neurons in human striatum. Parkinsonism Rel Disord
11:277–286.
Dawodu S, Thom M. 2005. Quantitative neuropathology of the entorhinal
cortex region in patients with hippocampal sclerosis and temporal lobe
epilepsy. Epilepsia 46:23–30.
Deaner RO, Isler K, Burkart J, van Schaik C. 2007. Overall brain size, and
not encephalization quotient, best predicts cognitive ability across
non-human primates. Brain Behav Evol 70:115–124.
Doetsch F. 2003. The glial identity of neural stem cells. Nat Neurosci
6:1127–1134.
Eriksson PS, Perfilieva E, Björk-Eriksson T, Alborn AM, Nordborg C, Peterson DA, Gage FH. 1998. Neurogenesis in the adult human hippocampus. Nat Med 4:1313–1317.
Fajardo C, Escobar MI, Buriticá E, Arteaga G, Umbarila J, Casanova MF,
Pimienta H. 2008. Von Economo neurons are present in the dorsolateral (dysgranular) prefrontal cortex of humans. Neurosci Lett 435:215–
218.
Frahm HD, Stephan H, Stephan M. 1982. Comparison of brain structure
volumes in Insectivora and Primates. I. Neocortex. J Hirnforsch 23:
375–389.
Gittins R, Harrison PJ. 2004. Neuronal density, size and shape in the
human anterior cingulate cortex: a comparison of Nissl and NeuN
staining. Brain Res Bull 63:155–160.
Grinberg L, Ferretti RE, Farfel JM, Leite R, Pasqualucci CA, Rosemberg S,
Nitrini R, Saldiva PHN, Jacob W. 2007. Brain bank of the Brazilian
aging brain study group—a milestone reached and more than 1,600
collected brains. Cell Tissue Bank 8:151–162.
Herculano-Houzel S, Lent R. 2005. Isotropic fractionator: a simple, rapid
method for the quantification of total cell and neuron numbers in the
brain. J Neurosci 25:2518 –2521.
Herculano-Houzel S, Mota B, Lent R. 2006. Cellular scaling rules for rodent
brains. Proc Natl Acad Sci U S A 103:12138 –12143.
Herculano-Houzel S, Collins C, Wong P, Kaas JH. 2007. Cellular scaling
rules for primate brains. Proc Natl Acad Sci U S A 104:3562–3567.
Jerison HJ. 1973. Evolution of the brain and intelligence. New York:
Academic Press.
Jorm AF, Jacomb PA. 1989. The informant questionnaire on cognitive
decline in the elderly (IQCODE): socio-demographic correlates, reliability, validity and some norms. Psychol Med 19:1015–1022.
Kandel ER, Schwartz JH, Jessel TM. 2000. Principles of neural science,
4th ed. New York: McGraw-Hill. p 19 –20.
Kumar SS, Buckmaster PS. 2007. Neuron-specific nuclear antigen NeuN is
not detectable in gerbil substantia nigra pars reticulata. Brain Res
1142:54 – 60.
Lange W. 1975. Cell number and cell density in the cerebellar cortex of
man and some other mammals. Cell Tissue Res 15:115–124.
Leigh SR. 1995. Socioecology and the ontogeny of sexual size dimorphism
in anthropoid primates. Am J Phys Anthropol 97:339 –356.
Lyck L, Dalmau I, Chemnitz J, Finsen B, Schroder HD. 2008. Immunohistochemical markers for quantitative studies of neurons and glia in
human neocortex. J Histochem Cytochem 56:201–221.
Marino L. 1998. A comparison of encephalization between odontocete
cetaceans and anthropoid primates. Brain Behav Evol 51:230 –238.
Morris JC. 1993. The CDR: current version and scoring rules. Neurology
43:2412–2413.
Mullen RJ, Buck CR, Smith AM. 1992. NeuN, a neuronal specific nuclear
protein in vertebrates. Development 116:201–211.
Nedergaard M, Ransom B, Goldman SA. 2003. New roles for astrocytes:
redefining the functional architecture of the brain. Trends Neurosci
26:523–530.
Nishiyama A, Yang Z, Butt A. 2005. What’s in a name? J Anat 207:687–
693.
Noctor SC, Martinez-Cerdeno V, Kriegstein AR. 2007. Contribution of
intermediate progenitor cells to cortical histogenesis. Arch Neurol
64:639 – 642.
Pakkenberg H. 1966. The number of nerve cells in the cerebral cortex of
man. J Comp Neurol 128:17–20.
Pakkenberg B, Gundersen HJG. 1988. Total number of neurons and glial
cells in human brain nuclei estimated by the disector and the fractionator. J Microsc 150:1–20.
Pakkenberg B, Gundersen HJ. 1997. Neocortical neuron number in humans: effect of sex and age. J Comp Neurol 384:312–320.
Pelvig DP, Pakkenberg H, Stark AK, Pakkenberg B. 2008. Neocortical glial
cell numbers in human brains. Neurobiol Aging 29:1754 –1762.
Reichenbach A. 1989. Glia:neuron index: review and hypothesis to account for different values in various mammals. Glia 2:71–77.
Rilling JK, Seligman RA. 2002. A quantitative morphometric comparative
analysis of the primate temporal lobe. J Hum Evol 42:505–533.
Roth G, Dicke U. 2005. Evolution of the brain and intelligence. Trends
Cogn Sci 9:250 –257.
Sarnat HB, Nochlin D, Born DE. 1998. Neuronal nuclear antigen (NeuN): a
marker of neuronal maturation in the early human fetal nervous system.
Brain Dev 20:88 –94.
Schoenemann PT, Sheehan MJ, Glotzer LD. 2005. Prefrontal white matter
volume is disproportionately larger in humans than in other primates.
Nat Neurosci 8:242–252.
Semendeferi K, Damasio H. 2000. The brain and its main anatomical
subdivisions in living hominoids using magnetic resonance imaging. J
Hum Evol 38:317–332.
The Journal of Comparative Neurology
THE HUMAN BRAIN AS A SCALED-UP PRIMATE BRAIN
Semendeferi K, Armstrong E, Schleicher A, Zilles K, Van Hoesen GW.
2001. Prefrontal cortex in humans and apes: a comparative study of
area 10. Am J Phys Anthropol 114:224.
Semendeferi K, Lu A, Schenker N, Damasio H. 2002. Humans and great
apes share a large frontal cortex. Nat Neurosci 5:272–276.
Shaham S. 2005. Glia–neuron interactions in nervous system function and
development. Curr Top Dev Biol 69:39 – 66.
Shariff GA. 1953. Cell counts in the primate cerebral cortex. J Comp
Neurol 98:381– 400.
Sherwood CC, Stimpson CD, Raghanti MA, Wildman DE, Uddin M, Grossman LI, Goodman M, Redmond JC, Bonar CJ, Erwin JM, Hof PR. 2006.
541
Evolution of increased glia–neuron ratios in the human frontal cortex.
Proc Natl Acad Sci U S A 103:13606 –13611.
Ullian EM, Sapperstein SK, Christopherson KS, Barres BA. 2001. Control
of synapse number by glia. Science 291:657– 660.
von Economo C, Koskinas GN. 1925. Die Cytoarchitektonik der Hirnrinde
des erwachsenen Menschen. Berlin: Springer.
Wolf HK, Buslei R, Schmidt-Kastner R, Shcmidt-Kastner PK, Pietsch T,
Wiestler OD, Bluhmke I. 1996. NeuN: a useful neuronal marker for
diagnostic histopathology. J Histochem Cytochem 44:1167–1171.
Zhang H, Miller RH. 1996. Density-dependent feedback inhibition of oligodendrocyte precursor expansion. J Neurosci 16:6886 – 6895.
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