The document summarizes research on how the integrity and tightness of the blood-brain barrier (BBB) is dynamically regulated in response to various physiological states and environmental factors throughout life. It discusses how the BBB develops and changes during key life stages like adolescence, pregnancy, and aging. The BBB undergoes remodeling during development, with tight junctions forming early in embryos, and pericyte and astrocyte interactions strengthening the barrier postnatally. Barrier properties also adjust during puberty due to sex hormone fluctuations, and pregnancy requires BBB adaptation to support vascular growth. Aging tends to increase BBB leakiness and susceptibility to breakdown.

1. Review
Blood–Brain Barrier Dynamics to Maintain
Brain Homeostasis
Marta Segarra,1,2,4,
* Maria R. Aburto,1,4
and Amparo Acker-Palmer1,2,3,
*
The blood–brain barrier (BBB) is a dynamic platform for exchange of substances
between the blood and the brain parenchyma, and it is an essential functional
gatekeeper for the central nervous system (CNS). While it is widely recognized
that BBB disruption is a hallmark of several neurovascular pathologies, an aspect
of the BBB that has received somewhat less attention is the dynamic modulation
of BBB tightness to maintain brain homeostasis in response to extrinsic
environmental factors and physiological changes. In this review, we summarize
how BBB integrity adjusts in critical stages along the life span, as well as how
BBB permeability can be altered by common stressors derived from nutritional
habits, environmental factors and psychological stress.
BBB Function
The vasculature of the CNS is a primary gateway regulating access of blood-borne molecules to
the brain, hence acting as an interface between the brain and peripheral influences. This physical
barrier, allowing only selective molecular transport across endothelial cells (ECs) and the brain
parenchyma, is known as the BBB [1,2]. The anatomical substrate of the BBB is the
neurovascular unit (NVU); a cellular assembly constituted by close functional association of
ECs, pericytes (embedded together with ECs in the basal lamina), and astrocytes, and supported
by other CNS cell types (Figure 1) [3]. ECs in the CNS seal the paracellular transport via junctional
protein complexes forming tight junctions (TJs) [4], allowing passage of nutrients and metabolites
only through tightly regulated transport and limiting the entrance of undesired products via efflux
transporters and restricted transcytosis [5]. Although the endothelium is central to the NVU, its func-
tion is regulated by input from adjacent cells such as pericytes, astrocytes, microglia, and neurons.
It is widely acknowledged that BBB dysfunction can contribute to the onset and detrimental out-
come of neurodegenerative disorders, cerebrovascular insults, and neuroinflammatory condi-
tions [6–8]. However, physiological variations of BBB tightness under healthy conditions, and
the mechanisms that regulate how BBB properties adapt to restore brain homeostasis are
less-well comprehended. In this context, this Review focuses on current knowledge on BBB dy-
namic adjustments in response to multifaceted physiological changes including those relating to
development, sleep/wake cycles, pregnancy, aging, diet alterations, and environmental
stressors. Exploring these dynamic regulations of the BBB may open new perspectives on under-
standing how systemic fluctuations can translate into different neuropathological states and pos-
sibly reveal new avenues for therapeutic options.
BBB Remodeling throughout Life
The BBB is a highly dynamic structure that adapts the exchange of molecules between the
bloodstream and the brain in response to homeostatic adjustments in health and disease.
Along the lifespan, BBB tightness adapts to continuous evolving changes in different physio-
logical states.
Highlights
The BBB is a dynamic structure that acts
as an active exchange platform to trans-
port molecules between the blood and
the CNS. The tightness and integrity of
the BBB vary in response to multiple fac-
tors, including environmental and sys-
temic influences. Regulation of BBB
integrity is crucial to maintain CNS
homeostasis.
Variations in the tightness of the BBB
have been documented in various con-
texts, including changes during develop-
ment, pregnancy and aging; in response
to environmental factors such as nutri-
tional state and extreme temperature;
and following psychosocial stress.
Aging, in particular, is a period suscepti-
ble to BBB breakdown, which can
contribute to neurodegeneration and
cognitive decline.
Maladaptation of the BBB to persisting
and/or severe stressors may contribute
to detrimental health outcomes.
1
Neuro and Vascular Guidance,
Buchmann Institute for Molecular Life
Sciences (BMLS) and Institute of Cell
Biology and Neuroscience, Max-von-
Laue-Strasse 15, D-60438, Frankfurt am
Main, Germany
2
Cardio-Pulmonary Institute (CPI), Max-
von-Laue-Strasse 15, D-60438,
Frankfurt am Main, Germany
3
Max Planck Institute for Brain Research,
Max-von-Laue-Strasse 4, 60438
Frankfurt am Main, Germany
4
These authors made an equal
contribution
*Correspondence:
Segarra@bio.uni-frankfurt.de
(M. Segarra) and
Acker-Palmer@bio.uni-frankfurt.de
(A. Acker-Palmer).
Trends in Neurosciences, May 2021, Vol. 44, No. 5 https://doi.org/10.1016/j.tins.2020.12.002 393
© 2020 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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2. Development
At embryonic stages, the fetus is primarily protected from harmful substances circulating in the
maternal blood by the placenta, which forms the blood–placenta barrier [9]. However, the BBB
is already formed at embryonic stages and confers additional protection to the developing
CNS. For detailed information on the molecular and cellular aspects of BBB development, we
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Figure 1. Multifactorial Modulation of Blood–Brain Barrier (BBB) Integrity. BBB tightness is accomplished by endothelial cells (ECs) tightly sealed by tight and
adherent junctions, which restrict the paracellular transport of blood-borne molecules. The ECs are surrounded by mural cells (pericytes or smooth muscle cells) that
are embedded in the basal lamina. Additionally, extensions of astrocytes (astrocytic end-feet) ensheath the vessels and mediate neurovascular communication.
Interactions between these different cell types at the neurovascular unit regulate BBB permeability. The tightness of the BBB and the crosstalk between its cellular
components are highly dynamic and modulated by intrinsic and extrinsic factors (schematically depicted in the circles). Fluctuations in BBB permeability occur in
physiological conditions. In addition, certain lifestyle habits, exposure to stressors and aging may favor BBB leakiness, through opening of tight junctions, increased
transcytosis, disrupted neurovascular communication, and neuroinflammation.
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3. direct readers to recent reviews [1,10]. In the classical view, the BBB was conceived as immature
in fetal and perinatal stages, but recent studies have challenged this concept, demonstrating that
the BBB is already functional in embryos, although barrier alterations could have a greater impact
in the developing, yet highly vulnerable brain [11,12]. In humans, it has been shown that important
barrier components are detectable as early as 12 weeks of gestation [13]. In rodents, the BBB is
formed around embryonic days 13.5–15.5 and crucially requires the recruitment of pericytes
[14,15], since pericyte–EC interaction is necessary for the establishment of TJs and the regulation
of transcytosis. The development of the BBB follows a spatiotemporal pattern [15] in synchrony
with other concomitant neurodevelopmental and vascularization processes, such as the forma-
tion of the intraneural vascular plexus or the cortical and tangential migration of neurons, guided
by intercellular crosstalk [3].
The complexity of the cellular organization of the NVU increases at postnatal stages with the
incorporation of astrocytes [16]. Defects in the assembly of astrocytic processes around the
vasculature lead to BBB leakage in young adult mice, despite unaltered pericyte coverage [17].
Both astrocytes and pericytes, which are regulators of the expression of molecules necessary
for BBB integrity, are required for barrier maintenance in adults [18,19]. BBB molecular compo-
nents differ between neonates and adults [20]: at perinatal stages, the expression of TJ proteins
(claudin-5 and occludin) and extracellular matrix components (laminin and collagen IV) is higher
than in adults, whereas expression of the pericytic marker platelet-derived growth factor receptor
(PDGFR)β increases gradually over time. These molecular differences between the developing
and adult NVU might account, for instance, for the well-preserved BBB integrity in neonatal
rats after acute stroke compared to vascular injury in adults [20]. Conversely, children and
young animals are more susceptible to BBB breakdown after infection [21,22]. One possible
explanation is that young animals might develop a more severe neutrophil-mediated acute
inflammatory response together with BBB breakdown compared to adults [22,23].
Adolescence, Pregnancy, and Reproductive Senescence
Puberty is a critical transitional period when the body, including the brain, undergoes a transfor-
mation towards adulthood. During puberty, there is an increase in luteinizing hormone and follicle-
stimulating hormone, triggering the production of sex steroids (testosterone and estradiol). Sex
hormones induce synaptogenesis and spine remodeling and modulate brain growth, potentiating
sexual dimorphism in some brain regions during adolescence [24–26]. The steroid hormones,
due to their small size and lipid solubility, can cross the BBB bidirectionally [27]. Although testos-
terone and estradiol treatments have been shown to decrease BBB permeability [28,29], little is
known about the effects of the peak of sex hormones on BBB integrity in adolescence.
Once sexual maturity is reached, in case of pregnancy, the BBB further adapts to the new phys-
iological conditions [30]. Pregnancy requires active vasculogenesis and angiogenesis, with in-
creased levels of circulatory vascular endothelial growth factor (VEGF) and placental growth
factor, both known for their ability to promote vascular permeability. In rats, potential effects of
such circulating factors on BBB integrity have been shown to be counterbalanced during preg-
nancy by the release of soluble VEGF receptor 1 in the maternal circulation, which prevents
BBB leakiness and brain edema during gestation [31]. In addition, pregnancy is also character-
ized by secretion of multiple hormones and cytokines that can potentially affect neuronal function.
Importantly, the BBB adapts to pregnancy by modulating the expression of influx and efflux trans-
porters, and preventing the passage of serum into the brain [30], which otherwise could trigger
seizures [32]. Indeed, efflux transporter inhibition induces rapid and sustained hippocampal sei-
zure activity preferentially in pregnant rats [33]. During pregnancy, the cerebrovascular system
is also exposed to hemodynamic fluctuations that can also affect BBB permeability [30,34], as
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4. denoted by BBB permeability increase following acute hypertension in pregnant rats. Moreover,
expression of the water channel aquaporin-4, localized at astrocytic end-feet, is increased during
pregnancy with a peak at mid-gestation. This increase has a potential link to edema formation in
response to pathological hypertension in pregnant dams [35].
During reproductive senescence, a decreased release of sex steroids and an increased secretion
of gonadotropins in the menopause/andropause have been reported [36]. There is a correlation
between the production of gonadotropins after ovariectomy and enhanced BBB permeability
[37]. When ovariectomy is performed in young adult and reproductive senescent rats, BBB
leakage can be mitigated by estradiol treatment in young animals. However, in aging rats,
which expose higher BBB disruption compared to young animals, the hormonal supplement is
ineffective [29]. Similarly, gonadal testosterone depletion increases BBB permeability in male
mice, and testosterone supplementation in castrated mice restores BBB integrity [28]. Mechanis-
tically, sex hormones and gonadotropins have been found to regulate the expression of TJs and
gap junctions (hemichannels connecting the plasma membrane of adjacent ECs) in different vas-
cular beds [28,37,38]. Also, estradiol treatment in ovariectomized rats increases the expression of
the glucose transporter, Glut-1, in brain ECs [39].
Aging
High-resolution magnetic resonance imaging in humans has revealed an age-dependent pro-
gressive increase of BBB permeability in the hippocampus, which was aggravated in individuals
with mild cognitive impairment, suggesting that increased BBB permeability in the hippocampus
might contribute to the development of dementia [40]. Relatedly, BBB breakdown is emerging as
an early biomarker for Alzheimer’s disease [41]. Increased soluble levels of the pericyte marker
PDGFRβ are found in the cerebrospinal fluid of aged individuals with BBB disruption [40], sug-
gesting that pericyte damage could be an effective biomarker for BBB breakdown. In line with
this observation, studies in pericyte-deficient mutant mice have demonstrated a progressive
age-dependent disruption of the BBB indicated by accumulation of plasma proteins and neuro-
toxic macromolecules in the brain parenchyma [42]. It has been also reported that the effective-
ness of the efflux transporter P-glycoprotein (P-gp) attenuates with age in healthy individuals
[43,44], especially in men [43]. A decline in P-gp function is also observed in Alzheimer’s disease
patients compared to age-matched healthy individuals [45], in line with the studies that connect
Alzheimer’s disease pathology with BBB dysfunction. Furthermore, aging influences the quality
of sleep (Box 1), and in mice, fragmented sleep has been shown to be associated with BBB
Box 1. BBB Tightness Is Modulated during Sleep
Circadian rhythms are recurring biological events with a period of ~24 h. BBB tightness is modulated by physiological cir-
cadian oscillations and sleep/wake phases [124,125]. Sleep is a vital physiological process that is regulated by the circa-
dian clock [126]. Sleep facilitates the clearance of macromolecules via the glymphatic system by enlarging the interstitial
space and favoring the convective flow between the interstitial and cerebrospinal fluids for an efficient removal of brain met-
abolic waste [127]. The transport of molecules through the BBB is also modulated by sleep. In rodents and flies, the activity
of the efflux transporter P-gp has been shown to decrease during sleep [128,129], suggesting that the permeability to xe-
nobiotics is more permissive during resting periods.
Chronic sleep deprivation affects multiple physiological processes such as glucose metabolism, neuronal function, and ce-
rebral blood flow that can ultimately contribute to diseases such as diabetes and Alzheimer’s disease [130–132]. Chronic
sleep disturbances in rodents result in decreased expression of TJ and increased paracellular permeability [132,133], in-
dicating that disruption of the BBB might be a mechanistic component in pathologies associated with sleep deprivation.
Moreover, in rats, disruption of rapid eye movement sleep has been shown to trigger BBB breakdown and increase
transcytosis in brain ECs [134]. Increased BBB permeability following sleep deprivation is associated with a mild pro-in-
flammatory status mediated by the release of cytokines, such as IL-1β, IL-6, IL-17A, and TNF-α, and increased levels
of proinflammatory molecules such as cyclo-oxygenase-2, NOS, or endothelin-1 [135,136].
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5. breakdown and extravasation of the proinflammatory cytokine tumor necrosis factor (TNF)-α into
the brain parenchyma [46]. Importantly, there is increasing evidence that sleep disturbances
might contribute to cognitive decline and development of Alzheimer’s disease [47,48].
Intriguingly, in a study using a heterochronic parabiosis model, systemic factors from the blood of
young animals rejuvenate age-related vascular and neurogenic degeneration in old animals [49].
This study identified growth differentiation factor (GDF)11 as one of the circulating factors that
could drive this rejuvenation effect. GDF11 does not cross the BBB but acts directly on the vas-
culature, inducing the release of the permeability factor VEGF [50]. This suggests that circulating
substances might impact on the brain endothelium and modulate BBB integrity in different stages
of the life span.
Nutritional and Metabolic Alterations and the BBB
Nutrition
Proper functioning of the CNS requires adequate nutritional support to sustain neuronal function.
Maintaining balanced levels of essential nutrients in the CNS environment requires that these nu-
trients are properly transported across the BBB. Macronutrients (e.g. glucose, fatty acids, and
amino acids) enter the brain by crossing the BBB from the circulating blood, mostly via carrier-
mediated transport or receptor-mediated transcytosis [51]. Moreover, some micronutrients [e.
g. niacin (vitamin B3), pyridoxine (vitamin B6), inositol, folate, and ascorbic acid (vitamin C)] are ac-
tively transported across the BBB and are maintained at higher concentrations in the brain com-
pared to other tissues. As a result, malfunction of these nutrients’ transport systems across the
BBB could lead to brain nutritional deficiencies despite of adequate whole-body levels. Further-
more, some of these micronutrients (e.g. ascorbic acid and folate) and certain ions (e.g. Na+
,
Cl−
and HCO3
−
) access the brain via the choroid plexus into the cerebrospinal fluid, constituting
a reservoir from which these nutrients slowly diffuse into the extracellular fluid, thereby making
the CNS concentrations resistant to whole-body depletion [52,53].
In animal models, malnutrition or dietary imbalances, for instance due to hypoproteic or high-fat
diets, have been shown to impair BBB function. These effects, however, were associated with
an inflammatory state, which makes it difficult to determine the direct effects of malnutrition on
BBB function [54].
It has been recently reported in mice that characteristic low transcytosis rates at the BBB require
the presence of Mfsd2a, a lipid flippase that transports phospholipids, including the essential fatty
acid docosahexaenoic acid (DHA), from the outer to the inner leaflet of brain EC plasma mem-
branes. This enrichment in DHA confers an increased fluidity to this inner leaflet, which prevents
the formation of transcytotic vesicles [55]. Low levels of DHA in the brain, due to either dietary def-
icits or an impaired transport of DHA by the fatty-acid-binding protein 5 into the brain, have been
shown to be associated with cognitive impairments and in Alzheimer’s disease mouse models
[56] and patients [57]. Thus, it would be interesting to determine whether BBB dysfunction as a
result of suboptimal DHA levels could be also contributing to these cognitive deficits.
Another interesting avenue in the context of nutrition and BBB is the ketogenic diet. In ketogenic
diets, carbohydrate consumption is drastically restricted, in favor of high fat and moderate protein
consumption. As a result, the body receives a minimal dietary source of glucose, which is re-
quired for all metabolic needs [58], thus, fatty acids become an major source of cellular energy
production. Fatty acids do not readily cross the BBB [59], but the liver metabolizes them into ke-
tones, which do cross the BBB, and in the absence of glucose, are the preferred source of energy
for the brain [60]. Ketones cross the BBB by passive diffusion or carrier-mediated transport
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6. facilitated by the monocarboxylate transporters, which are expressed in brain ECs, glia, and neu-
rons, which ultimately oxidize ketones to obtain ATP [61]. Ketogenic diet is known for its impor-
tant therapeutic effects in pathologies such as epilepsy and glucose transporter type 1 (Glut1)
deficiency – a metabolic disorder affecting the nervous system caused by poor glucose transport
across the BBB [60]. Ketogenic diet has been recently associated with increased cerebral blood
flow and P-gp transport through the BBB, which in turn could change the ecosystem of symbiotic
organisms in the gastrointestinal tract (i.e. ,gut microbiota) [62].
Gut–Brain Axis
Increasing evidence shows a bidirectional communication between the gut microbiota – the eco-
system of microorganisms residing in the gastrointestinal tract – and the brain, referred to as the
gut–brain axis (or the microbiota–gut–brain axis). The gut microbiota can signal to the brain via a
diverse set of pathways, including neural, endocrine, and immune signaling mechanisms, through
the production of bacterial metabolites such as short-chain fatty acids, branched chain amino
acids, and peptidoglycans [63]. It is increasingly recognized that the gut microbiota can influence
brain physiology and that alterations in the composition of this microbial ecosystem is involved in
the pathogenesis of a wide range of diseases. We next discuss briefly two studies that linked al-
terations in gut microbiota with BBB dysfunction [64,65]. In germ-free mice, a complete lack of
microbiota leads to a leaky BBB caused, at least in part, by a decrease in the TJ proteins occludin
and claudin-5 [64]. These mice present a leaky BBB already at embryonic day 16.5 (time when
the BBB is sealed), suggesting that the maternal microbiota is playing a fundamental role in de-
veloping a functional BBB. Importantly, this study identified that BBB integrity could be restored
by postnatal recolonization of the microbiota, implying a causal role for the microbiota in BBB
maturation [64]. Germ-free mice, however, represent a nonphysiological model, which compli-
cates the interpretation of studies based on them. As an alternative approach, antibiotics offer
a manipulation with greater temporal specificity, and have been used for instance to address
the importance of the gut microbiota in early life. Mice treated with a low dose of antibiotics
from embryonic stages to weaning, by administering these to the dam during pregnancy and lac-
tation, showed an increase in occludin and claudin-5 expression levels in hippocampus of males
and females. Moreover, males also showed an increase in occludin in the frontal cortex [65]. The
functionality of the BBB under these experimental conditions, and other manipulations of the gut
microbiome during early life, remains to be tested.
Metabolic Alterations
Obesity negatively impacts overall health. Adipose tissue is not only involved in energy storage,
but it is also an endocrine organ, as it secretes bioactive factors collectively known as cytokines
[66]. In obesity, secretion of these cytokines, which can be anti- or proinflammatory, is dysregu-
lated, as the expanding adipose tissue secretes large amounts of proinflammatory adipokines,
such as interleukin (IL)-6 and TNF-α, which can lead to the inflammatory responses and metabolic
dysfunctions associated with obesity [67]. Moreover, these adipokines can cross the BBB by a
saturable transport mechanism, although it is unclear whether obesity causes a change in the
transport of these adipokines through the BBB [68]. Obesity also leads to reduced transport
across the BBB of proteins involved in regulating appetite and food intake in the CNS, including
leptin and insulin [68]. Furthermore, a study in elderly people showed significantly elevated cere-
brospinal fluid/serum albumin ratio, a hallmark for BBB disruption, in obese subjects compared to
lean individuals [69]. Studies in animals have also been conclusive in demonstrating adverse ef-
fects of obesity on BBB function. Feeding high-saturated-fat diets to rodents (mice and rats),
has shown increased BBB permeability accompanied by changes in TJ proteins, such as
claudin-5, claudin-12, and occludin, as well as in cytoskeletal proteins including vimentin and tu-
bulin that are also essential for TJ stability [68]. However, as these studies did not specify the
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7. exact composition and nature of fat contents in high-fat diets, it remains difficult to draw mecha-
nistic conclusions. Furthermore, a proteomics study of brain microvessels from obese versus lean
mice identified several downregulated genes related to cellular energy metabolism, which could
be a cause of cerebral dysfunction in obesity [70]. In summary, inflammation in obesity might
be a cause for BBB disruption and enhancement of immune cell infiltration into the CNS.
Obesity also increases oxidative stress through different mechanisms such as hyperleptinemia,
low antioxidant defense, chronic inflammation, postprandial reactive oxygen species generation
[71], which may further contribute to obesity-mediated BBB disruption [68]. Conversely,
physical exercise has been shown to positively influence BBB integrity through a number of
anti-inflammatory and antioxidant effects [72].
Obesity and insulin resistance are often clustered together under the umbrella of metabolic
syndrome, which also includes high blood pressure and arterial stiffness, which can further
damage the BBB [73]. CNS insulin levels also depend on a functional BBB, as brain ECs regulate
insulin transport through the insulin receptor and via receptor-mediated transcytosis [74]. This
insulin transporter can be altered by physiological and pathological factors including hyperglycemia
and diabetic state. Moreover, diabetic insulin resistance is reported to accelerate cognitive decline
and increase dementia risk. In this context, studies have shown that compromised integrity of the
BBB precedes the associated cognitive dysfunction [41].
Environmental Stress and the BBB
Temperature
The BBB is sensitive to a broad spectrum of external or environmental influences. Extreme tem-
peratures have been reported to affect BBB permeability, and studies have shown an association
between hyperthermia and BBB disruption [75]. Hyperthermia is of particular interest due to its
deleterious effects on the CNS; a single episode of hyperthermia may cause short-to-long-term
neurological and cognitive dysfunction [76]. Hyperthermic stress could originate from an elevated
external temperature (e.g. heatstroke, the overheating of the body beyond40° C) as well as from
failure of the body’s thermoregulatory system (e.g. extreme fever), eventually leading to an eleva-
tion of brain temperature. Of note, most studies have been performed under conditions that in-
volve brain hyperthermia as a side effect, such as physical exercise in a warm environment
[77], opiate withdrawal [78], or methamphetamine intoxication [79,80]. Nonetheless, experiments
involving solely an elevation of brain temperature also showed BBB disruption and acute brain
edema [81]. A certain level of thermotolerance can develop in cells or tissues after repeated ep-
isodes of hyperthermia, through expression of heat-shock proteins. This phenomenon has
been ascribed to brain ECs in an in vitro BBB model, which could render a protective effect to
BBB disruption upon subsequent hyperthermic episodes [82]. On the other extreme, profound
brain hypothermia has also been shown to induce a mild BBB leakage and glial activation [81].
However, in rats, local hypothermia applied early after a stroke episode has been shown to be
neuroprotective, significantly reducing associated BBB disruption by reducing the loss of TJ
proteins [83].
Hypoxia
Among extrinsic insults known to induce BBB breakdown, hypoxia is probably the most charac-
terized, but many knowledge gaps remain. Hypoxia is among the endpoints of many disorders
such as stroke, cardiac arrest, respiratory distress, and carbon monoxide poisoning, but it can
also be caused by a reduction in the atmospheric oxygen partial pressure, for example, at high
altitudes. Hypoxia can disrupt the BBB and result in increased permeability, vasogenic edema,
and tissue damage [84]. Hypoxia induces expression of hypoxia-inducible factor-1; a major
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8. transcriptional regulator of the Vegf gene [85]. VEGF is expressed mainly by activated astrocytes
upon hypoxic stress [86] and it has been reported to induce changes in TJ proteins such as ZO-1
[87], claudin-5, and occludin [88,89]. VEGF also induces increased vascular permeability via
release of NO through endothelial NO synthase (NOS) activation [90]. Furthermore, hypoxia has
been associated with enhanced endothelial transcytosis as one major mechanism of BBB
opening. Several mediators, including NO, calcium influx, release of inflammatory cytokines, and
hemodynamic alterations, may be responsible for this alteration [91]. Moreover, hypoxia-induced
oxidative stress also contributes to BBB breakdown [92]. The different cell types at the NVU exhibit
distinct sensitivity to oxygen deprivation: brain ECs are markedly more sensitive than pericytes or
astrocytes, and pericytes more sensitive than astrocytes. Of note, the differential tolerance of
these cells correlates well with the oxygen levels they experience under physiological conditions
[84].
Hypobaric hypoxia after ascent to high altitude can lead to acute mountain sickness (AMS), which
develops within few hours after reaching high altitude. AMS includes headache, nausea, vomiting,
malaise, and can further progress into the more severe high-altitude cerebral edema, which can
be fatal [93]. While the pathophysiology of these high-altitude related syndromes is not fully
understood, it is recognized as multifactorial and involving underlying hypoxia and oxidative stress
[93,94].
Oxidative Stress
Oxidative stress is characterized by increased reactive oxygen species and reactive nitrogen
species. The high relative oxygen consumption by the brain (in humans, about 25% of the rest
of the body under resting conditions) enhances the brain’s susceptibility to oxidative stress.
Lifestyle, aging, external environmental factors, or individual genetic factors influence the degree
of oxidative stress in the CNS. To counteract this, the brain and its ECs are equipped with potent
defense systems against oxidative stress, including increased glutathione (GSH), glutathione
peroxidase, glutathione reductase, and catalase. GSH has been shown to play an important
role in maintenance of BBB integrity [95]. Furthermore, the healthy brain also contain antioxidants
such as superoxide dismutase and NF-erythroid 2-related factor 2 [95]. Brain ECs have a higher
mitochondria content, which is believed to fuel the high demand of energy-dependent transport
mechanisms and therefore to play an essential role in BBB maintenance [95]. However, it has
been also described that brain ECs obtain most of their energy from anaerobic glycolysis and
that mitochondrion’s main role is of a signaling organelle (e.g. for vascular tone regulation) [96].
In any case, the high demand of mitochondrial activity in brain ECs inevitably involves the
generation of an increased oxidative stress [95].
Psychosocial Stress and the BBB
Throughout life, individuals may face distinct types of psychological stress, varying in source,
duration, and severity. Exposure to different stressors activates a physiological response to
maintain homeostasis, and this adaptation to psychosocial stress can affect the brain in several
ways [97,98]. Typically, psychosocial stress activates the hypothalamic–pituitary–adrenal (HPA)
neuroendocrine axis through the release of corticotrophin-releasing hormone, leading to the
production of glucocorticoid hormones [98,99]. Acute rise of cortisol levels is beneficial to
generate efficient responses to threats in some circumstances; however, chronic exposure to
high levels of cortisol can be detrimental to brain health [100]. The integrity of the BBB is
modulated by neuroendocrine stimuli upon psychosocial stress, although the molecular and cel-
lular mechanisms controlling this process are still mostly unclear. In one study that examined
changes in BBB tightness upon psychosocial stress, long-lasting forced immobilization in
young rats induced opening of the BBB in correlation with increased levels of serotonin, although
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9. the direct effect of this hormone/neurotransmitter in BBB leakiness is unclear [101]. Short-term
immobilization is sufficient to cause albumin extravasation in certain areas of the rat brain, such
as the cerebellum, hippocampus, and hypothalamus [102]. Maternal separation in rats at perina-
tal ages induces BBB leakage in several areas of the brain, concurrent to increased levels of
serum corticosterone [103]. In humans, childhood psychological trauma is associated with in-
creased serum levels of the astrocytic protein S100β, a biomarker of BBB disruption [104]. In a
study in mice subjected to a forced swim test, the increased BBB permeability induced by psy-
chosocial stress was used to allow non-BBB-penetrant drugs to access the CNS and enhance
neuronal excitability [105]. It has been hypothesized that these stress-induced BBB alterations
could explain the enhanced CNS-associated side effects of certain peripherally acting drugs
[105].
The mechanisms linking psychosocial stress and BBB modulation have been examined in a num-
ber of recent studies. It has been shown for instance that in mice, HPA axis activation and ele-
vated corticosterone upon repeated social defeat induces the mobilization of bone-marrow-
derived monocytes and their recruitment into the brain by activated microglia, leading to a
neuroinflammatory response and alteration of brain vasculature, ultimately promoting anxiety-
like behavior [106–108]. Furthermore, acute psychosocial stress also has proinflammatory effects
mediated by activation of mast cells and is associated with BBB opening [109,110]. The hypoth-
esis emerging from these findings postulates that stress-induced neuroinflammation promotes
alterations at the NVU and modulates the integrity of the BBB [111]. Importantly, recent findings
suggest that the effects of psychosocial stress on BBB integrity contribute to variability in the sus-
ceptibility to stress [112]. Specifically, in this study, mice were exposed to chronic social defeat
stress (CSDS) and then stratified into two groups according to their social avoidance behavior;
that is, susceptible or resilient. The comparison between these behavioral outcomes revealed
that susceptible, but not resilient, mice displayed a significant reduction in TJ protein claudin-5
in the nucleus accumbens (NAc) [112]; a brain area involved in mood regulation [113]. This obser-
vation goes in line with morphological changes reported in endothelial TJs under restraint stress in
the prefrontal cortex, hippocampus, and amygdala of rats [114,115]. Of note, BBB permeability in
CSDS vulnerable mice was associated also with increased vascular transcytosis [112]. The au-
thors also showed that CSDS induced the recruitment of peripheral monocytes in the NAc and
leakage of the proinflammatory cytokine IL-6 in the parenchyma of stress-susceptible animals,
correlated with decreased expression of claudin-5. Moreover, reduction in claudin-5 expression
is correlated with depression-like behavior in mice and also with major depressive disorder in
humans [112], in line with clinical evidence from other studies of BBB disruption in neuropsychi-
atric patients [116–119]. In another study examining the relation between BBB breakdown and
social defeat stress, the authors found a specific reduction of the neuronal-derived cAMP in the
NAc in stress-susceptible mice [120]. BBB permeability, claudin-5 expression, and associated
depressive behavior were rescued upon treatment with cAMP. The glycoprotein reelin, recently
identified as an important modulator of BBB integrity [17], was found to be downregulated in
NAc of cAMP-deficient mice. Notably, injection of reelin within the NAc prevents IL-6 leakage in
the parenchyma of the NAc and ameliorates depression-like behavior in CSDS mice [120]. A
comparative transcriptomic analysis of NAc from stress resilient and susceptible mice showed
that the expression of the enzyme histone deacetylase (HDAC)1 was highly reduced in resilient
animals [121]. Of note, this study shows an epigenetic regulation of the expression of claudin-5
in psychosocial stress conditions and exposes a therapeutic target to promote resilience, since
HDAC1 pharmacological inhibition rescues the expression of the TJ protein claudin-5 and ame-
liorates social interaction in stress-defeated animals. Moreover, another transcriptomic study in
microglia derived from CSDS susceptible and resilient animals showed that microglia obtained
from stress-vulnerable mice displayed transcriptomic profiles highly enriched in pathways related
Trends in Neurosciences
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Trends in Neurosciences, May 2021, Vol. 44, No. 5 401

10. to inflammation, extracellular matrix remodeling, and phagocytosis [122]. This study confirmed
that BBB leakage was only observed in stress-susceptible, but not stress-resilient animals as
reported by others [112]. This suggests that leakage of blood-borne molecules into the brain
parenchyma upon BBB disruption triggered the activation of microglial cells in the CNS. In
fact, pharmacological depletion of microglial cells protected mice from the adverse effects of
CSDS [123]. Altogether, these findings support a key role for the brain–immune axis and
subsequent changes in BBB permeability in etiopathological mechanisms prompted by
psychosocial threats.
Concluding Remarks
BBB permeability is highly dynamic and responsive to multiple intrinsic and extrinsic signals to en-
sure brain homeostasis after alterations of different origins (Figure 1). The changes in BBB perme-
ability are generally driven by blood-borne substances, such as metabolites, hormones, or
cytokines, that either have a direct effect on the brain endothelium or induce a mild inflammatory
response that alters NVU function. Maladaptation to or chronicity of various insults can lead to a
range of pathologies. Therefore, prompt detection of changes in BBB integrity might constitute an
early diagnostic indication of such diseases [41], although considerable efforts are needed to
identify reliable biomarkers of BBB breakdown. Moreover, better understanding of the physiolog-
ical modulation of BBB properties is important to prevent deleterious consequences by means of
beneficial habits (see Outstanding Questions). Also, the consideration of possible BBB alterations
in pathological scenarios could help design novel therapeutic strategies, as well as optimize drug
administration practices. Specifically, this could include drug administration at suitable timing dur-
ing the circadian cycle, and stratification of drugs’ safety profiles according to age, metabolic co-
morbidities or stress. Particular attention is required to the risks associated with pharmacological
treatments during pregnancy and early childhood, especially in premature newborns devoid of
the protection of the placenta.
Acknowledgments
Work from the author’s laboratory cited in this review is supported by grants from the Deutsche Forschungsgemeinschaft
(SFB 834, SFB1080, SFB1193, FOR2325, EXC 115, EXC 147, EXC 2026) (A.A-P.), European Research Council
(ERC_AdG_Neurovessel Project Number: 669742) (A.A-P.) and the Max Planck Fellow Program (A.A-P.) and EU-CIG
293902 (M.S.).
Disclaimer Statement
The authors declare that they have no conflicts of interest in relation to the contents of this review.
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Multiple factors, such as diet, sleep,
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BBB. Often, some of these factors
coexist, for instance, during stressful life
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deprivation and a poor diet. Would it be
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Mild inflammation is often associated
with reduced BBB integrity, as
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psychosocial stress. Would anti-
inflammatory drugs be a suitable
treatment to prevent or revert BBB
leakage in these conditions?
The gut microbiome has been recently
shown to modulate BBB integrity. Since
the gut microbiota can be relatively
easily manipulated, could this be a
potential therapeutic target to regulate
BBB tightness? Moreover, many of the
above-mentioned conditions that have
a negative impact on BBB integrity are
also associated with disrupted gut
microbiome composition. What is the
specific contribution of gut microbiota to
alterations in BBB integrity observed in
such conditions?
Circadian rhythms modulate the opening
of the BBB. Should pharmacological
studies consider and explore the most
appropriate time windows for drug
administration? Could side effects
be diminished, or curative effects
enhanced, by adjusting treatment
timing? Additionally, BBB integrity is
differentially modulated as a function
of age. Should the risk assessment
of various drugs be stratified by age
groups?
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