The role of cytokines in modulating learning and memory and brain plasticity

1Brain plasticity is the ability of the brain to change its activity and modify its connections throughout life in response to extrinsic or intrinsic stimuli. Indeed, synaptic plasticity supports learning and memory processes during which the strength and efficacy of the synaptic transmission change between neurons, accompanied by structural modifications of spines, dendrites and axons. We will describe the role of cytokines in plasticity such as long-term potentiation (LTP), a key process for memory formation involving N-methyl-d-aspartate receptors (NMDAR) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPAR; ), and synaptic scaling, an operation by which a neuron adjusts the postsynaptic strength of all its synapses in order to maintain normal neuronal network functions. Neurons were, for a long time, thought to be the sole actors involved in plasticity but over the last decades, the roles of glial cells have switched from passive homeostatic elements to active modulators of synaptic plasticity and information processing. Among these glial cells are astrocytes that engulf synapses and surround blood vessels. Their many functions involve, among others, uptake and clearance of neurotransmitters such as glutamate and gamma aminobutyric acid (GABA;; ) and regulation of synaptic functions. Microglial cells also play a key role in modulating neuronal plasticity. They are a specialised population of tissue-resident macrophages broadly distributed in the brain parenchyma. Resting microglial cells are constantly scanning their environment by targeting synapses to monitor and regulate neuronal activity. In response to infections, stress, degenerative diseases or any changes in the nervous system both activated astrocytes and microglia produce cytokines that modulate a large variety of physiological and pathological processes.

2Cytokines are small pleiotropic signalling proteins classically secreted in response to pathogens or injury by cells of the immune system, including monocytes, macrophages, lymphocytes and vascular endothelial cells. This group of proteins comprises interleukins, chemokines, tumour necrosis factors, interferons, growth and cell stimulating factors and neurotrophins. In the brain, cytokines are constitutively expressed in various brain regions by activated glial and neuronal cells and are involved in several normal and pathological processes including sleep regulation, neuronal development, alteration of the blood–brain barrier, modulation of neurotransmitter metabolism and synaptic plasticity. There is an increased interest for the role of cytokines in the brain due to the presence of inflammation in the brain in a wide range of diseases such as Alzheimer’s disease (AD), major disorder depression, epilepsy, stroke, amyotrophic lateral sclerosis and arthritis. In this review, we will discuss how cytokines modulate learning and memory and plasticity in the normal central nervous system (CNS) function and under pathological inflammatory conditions. Learning and memory experiments provide a significant amount of data as they are an effective way of integrating molecular and cellular plasticity mechanisms with systems-level and behavioural changes. We will focus mainly on interleukin 1β (IL-1β), interleukin 6 (IL-6) and tumour necrosis factor α (TNF-α) as they are the most studied cytokines in the brain so far.

3Cytokines and their receptors are expressed in the brain by neurons, microglia and astrocytes and their involvement in structural changes at the synaptic level was reported over 20 years ago (for review,; ). Periodic changes in synaptic transmission that underlie modification of behavioural states are often associated with adjustments in neurotransmitters at the synapse and those are influenced by cytokines. An overview of the effects of each cytokine on plasticity and learning and memory are summarised in Figure 1 and the mechanisms of action are presented in Figure 2.

4Under normal resting conditions, IL-1β levels are usually below the level of detection and a stimulus is necessary to augment IL-1β expression. Many reports support the role of IL-1β in spatial recognition and contextual learning. IL-1β messenger RNA (mRNA) expression was found to be induced following fear conditioning and during spatial memory tasks. No impairment was detected in IL-1 receptor (IL-1r) knockout (KO) mice for auditory cued memory nor visual memory and intrahippocampal injection of IL-1β after training impaired the consolidation of contextual but not auditory fear conditioning memory. However, mice depleted of any endogenous IL-1 receptor displayed hippocampus-dependent learning deficits, which highlight the complexity of IL-1 function in the brain (Table 1) and point to a possible role of IL-1β and IL-1α in normal memory function. A few studies have also shown that IL-1β is induced during short- and long-term plasticity and is necessary for LTP maintenance. It is important to stress that IL-1 effects on memory are dose- and age-dependent (see Table 1): at low concentrations, injections of IL-1β appeared to promote synaptic plasticity and improved performance in passive avoidance conditioning whereas injections of higher IL-1β doses following a learning task resulted in impaired memory. Most reports use ‘adult’ mice, usually between 2- and 4-month-old; however, an interesting study points out that the role of IL-1 might change when animals get older as only young (3 months) but not older mice (6 months) display deficits in spatial recognition. The authors suggest that synaptic sensitivity to IL-1β and that IL-1β function might evolve with age and with the appearance of age-related brain inflammation and elevation of proinflammatory molecules.

5Peripheral IL-1β injection stimulates the release of monoamines in the brain, which are known to modulate learning and memory (for review, ); for example, passive avoidance and acquisition of the water maze task were impaired in rats treated with a norepinephrine agonist; spatial memory performance was impaired in rats treated with 5-HT receptor agonist and in rats having depleted dopamine. IL-1β can also modulate plasticity and memory by promoting the production of neurotrophic factors: nerve growth factor (NGF) stimulates acetylcholinesterase activity and facilitates LTP induction; brain-derived neurotrophic factor (BDNF) activates multiple signalling pathways involved in synaptic plasticity and memory formation such as TrkB–ERK pathway that leads to glutamate and GABA release, potentiation of NMDAR and upregulation of membrane AMPAR (reviewed in the study by ).

6Rodent studies have shed light on the role of IL-6 in learning and memory (Figure 1) in different brain regions. IL-6 KO mice showed deficits in spatial learning tasks and when IL-6 was blocked via the injection of anti-IL-6 antibodies in orbitofrontal cortex (OFC), rats had deficits in a reversal learning task. IL-6 role in the striatum was investigated by Brennan et al; they showed that an intraperitoneal (IP) injection of IL-6 prior to an escape/avoidance task did not have any effect on the performance of the animal. This task involves the dorsal striatal learning system and it is possible that IL-6 might not be required in that instance, or that the cytokine concentration was too low to have an effect. Several publications have also described a role of IL-6 in the hippocampus. Hippocampal-dependent avoidance learning performance was impaired when IL-6 was injected in the hippocampus. Young and adult IL-6 KO mice perform better in a radial maze task compared to their wild-type (WT) counterparts; this difference was particularly noticeable in young mice, which suggest that the chronic lack of IL-6 facilitates learning and memory processes at a juvenile stage. IL-6 mRNA levels were upregulated after LTP induction in vitro and in freely moving rats, and blocking IL-6 led to prolonged LTP at the perforant path and improved memory in a Y-maze task. Overall, these results suggest that IL-6 production and function in memory and plasticity might vary with age and brain region (Table 2).

7The mechanisms by which IL-6 modulates cognitive function are far from being elucidated (Figure 2). Decreased ERK1/2 activation after IL-6 application correlates with impairments of LTP. A hypothesis is that the memory facilitation observed in mice lacking IL-6 could be due to the absence of inhibition from the endogenous opioid system as µ-opioid receptors were downregulated in IL-6 KO mice and therefore unable to dampen learning and memory. IL-6 is also known to interact with the cholinergic system and peripheral injection of this cytokine reduced scopolamine-induced impairment of a passive avoidance task. Moreover, intracellular calcium response to NMDA or glutamate was enhanced following chronic treatment of cultured cerebellar-granule neurons with IL-6 meaning that IL-6 can also potentiate neurotransmitter responses and thus any dependent behaviour.

8Very few studies have investigated TNF-α role under physiological conditions. Mice lacking TNF-α performed poorly in a novel object recognition task and in the Barnes maze test that evaluates spatial memory and learning effectiveness. A positive effect of TNF-α on learning and memory was also described in an escape/avoidance task where rats injected with TNF-α (6 μg/kg, intraperitoneal) made more avoidance responses and fewer escape responses compared to controls.

9TNF-α role in synaptic activity has been more extensively studied. TNF-α regulates synaptic scaling by promoting the insertion of AMPAR lacking GluR2 subunit at the plasma membrane of synapses, hereby promoting excitatory signalling over inhibitory signalling. TNF-α also modulates gliotransmission by elevating astrocytic calcium levels which in turn increase by twofold the number of glutamatergic exocytic vesicles. Released glutamate activates neuronal NMDAR at granule cells synapses of the dentate gyrus, hereby increasing excitatory synaptic activity and potentiation of GC synapses.

10Beattie et al. showed, using neuronal cultures and acute hippocampal slices, that TNF-α was required for preservation of synaptic strength and increasing the number of AMPAR at the membrane, the number of synapses and the frequency of miniature excitatory post-synaptic currents. Concomitant to these changes, TNF-α was found responsible for an increase in spine size of dendritic branches with recent spine loss, which constitute another synaptic scaling feature. Other studies have confirmed that constitutive TNF-α is necessary for maintaining AMPAR at the synapse, and described how TNF-α also triggered GABAA receptors internalisation, highlighting the unique ability of this cytokine to modulate both excitatory and inhibitory plasticity. Even though TNF-α can be produced by neurons, the above effects were generated by glial TNF-α in response to extracellular glutamate.

11The role of TNF-α in other forms of plasticity is unclear. Some studies reported that LTP induction or maintenance did not require TNF-α as TNF-α KO mice displayed normal LTP in the CA1 pyramidal layer. However, a recent report highlighted a significant TNF-α-dependent inhibition of LTP in the stratum radiatum but not the stratum oriens of rats after either high-frequency priming stimulation or treatment with TNF-α (1.8 nM) via the phosphorylation of p38-MAPK, ERK and JNK, pointing to a differential regulation of LTP in the CA1 basal and apical dendrites. Findings regarding TNF-α and LTD are also conflicted. Although Albensi et al. showed that TNF-α is a key factor for the induction of hippocampal LTD via nuclear factor kappa B (NFκB) activation, that was not the case in other laboratories. Overall, these varying results could be explained by different experimental conditions such as theta burst or high-frequency stimulation, the area in which the stimulation and recordings are made, the use of different TNF-α concentration, time of incubation, cell culture or brain slices.

12Central and peripheral inflammation in response to pathogens, injury or disease is a defence mechanism by which microglia and astrocytes become activated and potentially produce chemoattractant proteins that promote extravasation of monocytes from the periphery into the brain. Therefore, in the inflamed brain, cytokines are produced by microglia, astrocytes, neurons, peripheral inflammatory cells, endothelial cells, pericytes and choroid plexus; and cytokine receptors are present on neurons, astrocytes, microglia and vascular endothelial and perivascular cells. TNF-α, IL-1β and IL-6 have the ability to stimulate each other’s production and can act synergistically to modulate neuronal responses, plasticity and learning and memory (Figure 2), often translated by an elevated excitability of the brain and learning and memory changes that we describe below.

13A detrimental role of IL-1β on learning and memory was described in an inflammatory environment (Figure 1) created by generating chronic elevated IL-1 levels in the brain. Spatial memory deficits were reported after chronic injection of IL-1β into the lateral ventricles of rats in an eight-arm radial maze. Mouse models of inflammation using chronic hippocampal overexpression of IL-1β for 2 weeks led to the appearance of inflammatory markers such as activated glia, elevated prostaglandin E2 (PGE2), increased hippocampal proinflammatory cytokine and chemokine mRNAs and lower levels of the plasticity-related gene Arc. These transgenic mice displayed delayed acquisition and decreased retention in the spatial water maze task, and impaired long-term contextual fear memory, while hippocampal-independent and short-term memory remained intact.

14Models of peripheral infections have also been used to study cytokines in the brain. Peripheral infection can cause significant impairment of cognitive functions in individuals already suffering from neurodegenerative disease (Table 3; ) and to a lesser extent in healthy individuals. When injected intraperitoneally with Escherichia coli, IL-1β is upregulated in the aged rat hippocampus. The involvement of IL-1β in learning and memory was also studied in mice injected with Legionella pneumophila before subjecting them to the Morris water maze (MWM). Sick mice displayed impaired learning; their task performance was, however, restored to control levels if they were treated with an anti-IL-1β antibody. It is worth noticing that the effects of IL-1β depend (1) on the intensity of the protocol, as a high frequency of training sessions were more likely to highlight an IL-1β–related memory impairment in rodents than when spaced protocols were used and (2) on the concentration of IL-1β that is injected in the animal, with low dose (100 ng/mouse) leading to learning impairment and high dose of IL-1β (1000 ng/mouse) facilitating spatial navigation learning. Lipopolysaccharide (LPS) is another compound regularly used to study the effects of cytokines during an infection and it classically generates sickness behaviour-related symptoms. Following LPS injection in mice lacking caspase-1, the enzyme that cleaves the inactive pro-IL-1 into its active form, mice displayed reduced sickness behaviours and decreased levels of proinflammatory markers such as PGE2. IP injection of LPS increased IL-1β protein levels in the cortex, hippocampus and hypothalamus of rat and impaired contextual but not auditory fear conditioning, an effect that was abolished by subcutaneous injection of the endogenous antagonist IL-1ra. A mouse model of septic encephalopathy with blood–brain barrier disruption was characterised by astrogliosis and upregulation of hippocampal IL-1β and its receptor IL-1R1, and altered LTP was recovered when hippocampal slices were pre-incubated with an IL-1R1 antagonist. IL-1β impairment of LTP was recorded in the Schaffer collateral–CA1 synapses and in the associational/commissural fibre–CA3 synapses; LTP deficit was found to be dependent on the activation of mitogen-activated protein kinases (MAPKs) and the presence of NMDAR. Interestingly, LTP was not impaired at mossy fibre–CA3 synapses, possibly because these synapses are NMDAR-independent.

15Stressors like inescapable shocks also trigger an increase of IL-1β protein levels in the brain, although this increase can be difficult to highlight as it is likely to be masked by an inhibitory action of glucocorticoids. Chronic mild stress triggered an increase of hippocampal IL-1β protein levels and depression-like symptoms in rodents, but mice lacking the IL-1 receptor or with an astrocytic overexpression of an IL-1 antagonist did not display stress-induced behavioural nor neuroendocrine changes. Social isolation also elevated Il-1β protein levels in the hippocampus and the cerebral cortex and consequently led to contextual fear-conditioning impairment. Similar to isolation studies, rats infused with IL-1β in the hippocampus displayed contextual but not auditory deficits after fear conditioning, and this effect was prevented by intracerebral injection of IL-1ra. IL-1 was also found to impair working memory using a three-panel runway task. In a rodent model of seizure where the mouse cecum is ligated then punctured, IL-1β mRNA expression in the hippocampus was upregulated, escape latency in the MWM was impaired and LTP decreased; the use of an IL-1 antagonist rescued LTP and learning and memory abilities. High hippocampal IL-1β levels found in aged or stressed rats was also associated with impaired LTP. Under stress conditions, it is likely that microglial IL-1β production increase is linked to the activation of the purinergic receptor P2R×7, which is a key protein that supports stress and depression mechanisms.

16In a model of Parkinson’s disease, elevated IL-1β production in the brain worsened neurodegeneration course by supporting neuronal death, either directly or via nitric oxide production. IL-1β levels are elevated in AD brain as IL-1β was produced in response to Aβ exposure in cell culture and was shown to contribute to cognitive deficits, tau phosphorylation and Aβ pathology in aged 3× Tg-AD mice. Aβ-induced astrocytic IL-1β is also suspected of supporting the overexpression and activity of acetylcholinesterase hence exacerbating the cholinergic dysfunction linked to cognitive impairment observed in AD.

17Some mechanisms by which IL-1β modulates memory function have been described. It has been reported that the synaptic sensitivity to IL-1β increases with age due to a ratio change of the IL-1-receptor type 1 subunits AcP (proinflammatory) and AcPb (prosurvival) that facilitates memory impairment. There is evidence also for the arachidonic cascade and the glutamatergic system to be influenced by IL-1β. Bilateral injection of IL-1β in the hippocampus of rats increased cyclooxygenase-2 (COX2) protein levels and activated the arachidonic cascade and the release of PGE2 in the hippocampus. Hippocampal PGE2 injection led to more working memory errors than in control group. Similarly, selective and competitive NMDAR antagonists caused deficits in working memory performance in the three-panel runway task. An NMDAR agonist concurrently administered with IL-1β into the hippocampus reversed the working memory impairment induced by the cytokine. IL-1β also modulates calcium influx through NMDARs. Moreover, elevated IL-1β protein levels in the brain could inhibit LTP by interfering with BDNF signal transduction and dendritic spine morphology.

18In illnesses such as depression, arthritis and others where chronic inflammation is settled, patients report sickness and depressive behaviours and cognitive decline, often accompanied by elevated levels of IL-6. Individuals given a dose of LPS did not report feeling sick but performed poorly during declarative memory tests and better in working memory tasks. A lot of rodent studies have confirmed these detrimental effects of IL-6 on cognition and behaviour. Mice lacking IL-6 displayed reduced sickness behaviour and depressive-like social behaviour induced by systemic and central injections of IL-1β and LPS was lower in IL-6−/− mice than in their WT counterparts. Similarly, turpentine-induced abscess or influenza infection did not cause any behavioural deficits in IL-6−/− mice. Sparkman et al. evaluated the ability of these animals to integrate new information after the acquisition period of an MWM task by injecting LPS. Contrary to WT mice, IL-6 KO animals failed to display any working memory impairment and lacked the expected LPS-induced increase in TNF-α and IL-1β mRNA in hippocampal neurons but not in the periphery. However, LPS injection still activated a marker of neuronal activity c-Fos in neurons of the nucleus tractus solitarius that receive peripheral information via the vagal nerve. These data support the fact that IL-6 is not fundamental for periphery-to-brain communication but is required for LPS-induced production of TNF-α and IL-1β in the brain and for the expression of some behavioural impairments. Mice lacking or expressing IL-6 have also been used to highlight the role of IL-6 in the limbic system in relation to anxiety-like and depressive-like behaviours. IL-6 KO mice showed alterations in such behaviours. Decreased exploratory behaviour in a light–dark transfer test and digging behaviours were reported in mice overexpressing IL-6 together with a longer time spent immobile in a forced-swim test (; Table 2) as a readout of the ability of the animal to adapt upon exposure to an inescapable stressor.

19IL-6 plays a key role in activating astrocytes and microglia during inflammatory conditions. IL-6 KO mice displayed reduced reactive astrogliosis and microgliosis after IP injection of the toxic glutamate receptor agonist kainic acid (model of seizure; ) or subcutaneous injection of the neurotoxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (model of selective dopaminergic cell injury, which occurs in Parkinson’s disease; ). Indeed, IL-6 KO mice had a reduced astrocytic population and a decreased ability to activate microglia; and in a murine model of experimental autoimmune encephalomyelitis, IL-6 blocked the astrocytic response to interferon-γ and the release of microglial inhibitory molecules such as galectin-1 and HO-1. This glial cell activation by IL-6 is likely to happen in ageing and neurodegenerative diseases. Studies have described the elevation of IL-6 in the plasma of elderly individuals and its correlation with cognitive decline, with females being more sensitive to higher IL-6 levels (Table 3;;; ). These results were confirmed in a mouse model of accelerated ageing in which IL-6 protein levels were increased by 50% and 30% in the hippocampus and cortex of aged mice, respectively, compared to controls. Similarly, IL-6 gene and protein expression in glial cells were found to be upregulated in the cerebellum, cortex and hippocampus of mice as they age naturally. IL-6 detrimental effects on cognition could be linked to neurodegeneration and calcium homeostasis which is perturbed in the brain of Alzheimer’s patients and aged individuals, with the expression of a key calcium-binding and buffering protein calbindin being reduced in rodent models of dementia (;; Figure 2). Mice chronically expressing astrocytic IL-6 were found to express a progressive age-related decline in avoidance learning performance that correlated with pre-synaptic loss, typically associated with behavioural deficits and a decrease in cortical and hippocampal neuronal calbindin that plays a protective role in AD. However, in the early phase of AD, IL-6 participates in a reduction in Aβ deposition as IL-6 overexpression in a mouse model of AD led to enhanced microglia activation and phagocytosis activity.

20The role of TNF-α in learning and memory under inflammatory conditions is likely to be age dependent. Indeed, studies using 30-day-old mice overexpressing TNF-α did not find any learning and memory impairment in the water maze test, whereas older mice showed impaired memory in a passive avoidance task, in the water maze and in a three-panel runway task. Adult rats chronically overexpressing five times the normal levels of murine neuronal TNF-α also display spatial memory impairments in a water maze paradigm. In these animals, hippocampal synaptic plasticity measured by LTP and paired-pulse facilitation are comparable to those measured in the immature hippocampus of neonates, which support a role of TNF-α in maturation of the synaptic network. Another report points to a beneficial role of TNF-α in memory recovery as mice lacking TNF-α that have recovered from pneumococcal meningitis displayed impaired water maze performance compared to WT controls.

21Chronic pain is often accompanied by memory deficits and a reduced hippocampal volume (Table 3;; ). In a model of chronic pain, a pared sciatic nerve leads to microglia activation, increases TNF-α levels and impairs LTP in the hippocampus, reduces the density of presynaptic boutons, as well as the functional synaptic connectivity and BDNF expression in CA1 neurons. These synaptic changes are TNF-α-dependent as similar results were obtained when TNF-α was injected in the brain, but not when the experiment was repeated with mice lacking the TNF-α receptor TNFR1.

22Astrocytic and microglial TNF-α release under inflammatory conditions is a feature of neurodegenerative diseases and is associated with increased levels of Aβ, tau and neuronal cell death. CSF levels of TNF-α, produced by brain cells, are increased in some AD patients; TNF-α mediates LTP inhibition as well as memory deficits caused by Aβ. In aged APP/PS1 mice, a model of AD in which TNF-α levels are twofold higher than in their WT counterparts, there is a marked TNF-α-dependent inhibition of LTP possibly due to elevated TNF-α production by Aβ-dependent glial cell activation. Several other studies have reported that in mice and hippocampal slices, TNF-α abolished LTP and had deleterious effects on working memory, while cognitive deficit is reduced by pharmacological inhibition of TNF-α in mice. Interestingly, earlier in vitro studies have reported a protective role of TNF-α in AD, with human TNF-α levels inversely correlated with levels of a marker of neuronal degradation tau and in vitro data linking TNF-α incubation of neuronal cells to an increase in bcl-2, which can downregulate apoptosis. TNF-α has also been shown to protect against glutamate, free radical and Aβ toxicity in enriched cultures of primary neurons.

23In an inflammatory context, TNF-α levels can rise from picomolar to millimolar levels; phosphorylation of MAPKs (Figure 2) have been described during TNF-α inhibition of LTP in the molecular layer of the dentate gyrus and hippocampal apical dendrites. Elevated TNF-α concentration can also activate NFκB and potentiate NMDAR and AMPAR via TNFR1 activation, blocking glutamate transporter activity and promoting glutamate neurotoxicity, leading eventually to neuronal and oligodendrocyte cell death. Moreover, TNF-α favours synaptic transfer of calcium-permeable AMPAR lacking GluR2 subunit, hence potentially increasing the risk of neuronal toxicity.

24Surprisingly, the effects described above do not occur in the dorsolateral striatum of rodents. In this region rich in inhibitory GABAergic neurons, TNF-α promotes the internalisation of AMPAR permeable to calcium thereby reducing synaptic strength, which could be an adaptative mechanism of the brain to delay motor symptoms in diseases characterised by movement disorders and elevated TNF-α levels. Overall TNF-α roles on plasticity depend on the anatomical location of TNF-α action and on the level of expression of TNF-α.

25Cytokines are much more than basic signalling inflammatory molecules as they have key roles in modulating cognitive functions in physiological conditions. We propose that the length, strength and location in a particular area of the brain of cytokine activity will dictate the activation of specific signalling pathways together with the stimulation of other inflammatory systems that will contribute to the complexity of the response. This renders the distinction between mechanisms that would be solely engaged either in physiological or in inflammatory conditions very difficult. We suggest that the homeostasis equilibrium shifts when pathological, chronic expression of cytokines sustains in time; then the cytokine actions often become detrimental and lead to neuronal death and learning and memory impairments. In that view, a better understanding of the interaction between the whole immune system and synaptic plasticity would certainly be key to develop treatments for neuronal, cognitive and mood disorders that present deficits in these mechanisms.