Astroglia Abnormalities in Post-stroke Mood Disorders

Overview of Stroke

Stroke is the leading cause of serious, long-term disability in the USA. Stroke can be categorized into two different types—ischemic stroke and hemorrhagic stroke. According to a recent report from American Heart Association (AHA), of all the strokes in the USA, 87% are considered ischemic stroke, 10% are intracerebral hemorrhage (ICH), and 3% are subarachnoid hemorrhage (SAH) (Virani et al. 2020).

Ischemic stroke occurs when the brain’s blood vessels become narrowed or blocked (Moskowitz et al. 2010). This is problematic for the brain because it depends on arteries to carry fresh blood to it in order to bring in oxygen and glucose (as well as other nutrients) and remove carbon dioxide and cellular waste. If the arteries are blocked for too long, neurons, the most important cell type in central nervous system (CNS), cannot produce enough energy and will stop functioning and die. Ischemic stroke is most often caused by atherosclerosis, which is the buildup of fats, cholesterol, calcium, and other substances called plaque on artery walls (Campbell et al. 2019; Moskowitz et al. 2010; Lo et al. 2003). Atherosclerosis is a progressive disease and begins with damage to the inner layer of an artery. Over time, the plaque will harden, and arteries will become narrower, limiting blood flow and eventually forming blood clots, further narrowing arteries and limiting the flow of oxygen-rich blood. A blood clot may completely block blood flow or break apart and trigger a stroke. Although the cause of atherosclerosis is not known, factors such as high blood pressure, high cholesterol, obesity, diabetes, and tobacco may increase risk for the disease.

Ischemic stroke can be further categorized into thrombotic stroke and embolic stroke (Barber and Demchuk 2003; Dirnagl et al. 1999). Thrombotic stroke, or cerebral thrombosis, is caused by a blood clot (thrombus) that blocks blood flow in a damaged cerebral artery leading to or within the brain. Large-vessel thrombosis occurs when arteries such as the carotid or middle cerebral are blocked, and small-vessel thrombosis occurs when the brain’s smaller, yet deeper, penetrating arteries are blocked. Embolic stroke, or cerebral embolism, is caused by a wandering blood clot (embolus) in an artery somewhere other than the brain such as the neck or heart. The bloodstream carries the clot, which blocks a blood vessel in or leading to the brain (embolism). Again, this restricts the flow of blood to the brain and results in near-immediate physical and neurological problems. The main cause of embolism is atrial fibrillation, an irregular heartbeat which has the potential to develop blood clots within the heart. These clots can then circulate to the brain, causing ischemic stroke. The risk of stroke from atrial fibrillation depends on many factors including age, high blood pressure, and diabetes.

Hemorrhagic stroke, on the other hand, is caused by the leaking or bursting of blood vessels (Moskowitz et al. 2010). The leaked blood from vessels in the brain puts pressure on brain cells and damages them. Hemorrhagic strokes can be a result of conditions affecting blood vessels, including high blood pressure, overtreatment of blood thinners, aneurysms, trauma, cerebral amyloid angiopathy (protein deposits in blood vessel walls that weaken vessel walls), and arteriovenous malformation (the rupture of an abnormal tangle of thin-walled blood vessels). Intracerebral hemorrhage is the most common type of hemorrhagic stroke, occurring when brain arteries burst and flood surrounding tissue with blood. Subarachnoid hemorrhage refers to bleeding in the subarachnoid space, which is the area between the brain and tissues that surround the brain.

There are a number of detectable clinical symptoms and pre-stroke warning signs (Saengsuwan et al. 2017; Mochari-Greenberger et al. 2014) including sudden unilateral weakness (paralysis of face, arm, or leg on one side of the body), sudden trouble with speaking and understanding speech, sudden trouble with walking, loss of balance, sudden blackened or blurred vision in one or both eyes, double vision, and sudden and severe headaches perhaps accompanied by vomiting, dizziness, or altered consciousness. These symptoms are dependent on which side and region of the brain are affected and how severely the brain is damaged. It is often the case that only one side of the body is affected.

After a stroke, some patients will recover fully, others will be disabled long term or for the rest of their lives, and some will die if the damage to their brain is too severe. Approximately 3% of males and 2% of females reported that they were disabled because of stroke (Virani et al. 2020). Some problems that patients may experience after stroke can be managed by medical professionals through different types of rehabilitation therapy (Campbell et al. 2019). Regaining the activities of daily living become the focus of rehabilitation after stroke. For stroke survivors, the first 3 months post stroke are the most important period for recovery and when patients show the most improvement (Grefkes and Fink 2020; Ronning and Guldvog 1998; Poulin et al. 2016). The goal of rehabilitation is to restore function as close as possible to “normal” or find strategies to work around new limitations. For example, speech therapy can help those who have trouble understanding or producing speech. Physical therapy helps with relearning movement and coordination skills lost after stroke which may include paralysis and/or weakness on one or both sides of the body. Occupational therapy improves daily activities such as eating, drinking, dressing, bathing, reading, and writing. Stroke patients may have trouble with chewing and swallowing or bladder and bowel control, so occupational therapists may also help them regain these skills. Even though rehabilitation psychologists and neuropsychologists can help, many patients still suffer long-term complications (effects). These complications are not only physical symptoms such as weakness, paralysis, difficulty swallowing, gait instability, and falls and fractures but also pain syndromes, depression, anxiety, pseudobulbar affect, epilepsy, cognitive impairment, and dementia (Virani et al. 2020).

Stroke has a large impact on public health and significant financial costs. A comprehensive study was conducted on 97,374 hospitalizations with a primary or secondary diagnosis of stroke from 2006 to 2008. It found that ischemic, hemorrhagic, and other types of strokes had average hospitalization costs of $18,963, $32,035, and $19,248, respectively. Overall, hemorrhagic stroke cost $14,499 more than ischemic stroke. In addition, the direct medical cost of stroke in 2008 in the USA was estimated to be $18.8 billion, and for the same year, the estimated per-person expenditure for stroke care was $7657 (Wang et al. 2014a). These costs largely increased based on the 2020 update of Heart Disease and Stroke Statistics from America Heart Association (AHA). According to the update, the average annual direct medical cost of stroke in the USA was $28.0 billion and direct care was estimated at $7902 in 2014 and 2015 (Virani et al. 2020). The costs of hospitalizations involving stroke are high and vary greatly by type of stroke, diagnosis status, and comorbidities.

In this chapter, before briefly describing the pathological processes of ischemic stroke, we will focus on the role of astrocytes in post-stroke mood disorders.

Pathological Processes of Ischemic Stroke

Acute Responses and Cell Death After Ischemic Stroke

Healthy brain tissue has a high level of consumption of oxygen and glucose. Neuronal function relies on continuous ATP production which requires oxygen and glucose from the blood. When a focal ischemia stroke takes place, cerebral blood flow is less than 20% of the normal standard at the ischemic core (IC) (Dirnagl et al. 1999; Lo et al. 2003). Impairment of cerebral blood flow restricts delivery of glucose and oxygen, impairing the energetics required for ion gradients and leading to a loss of membrane potential in glia and neurons.

In focal ischemic stroke (FIS), the regions determine the pattern of cell death. Necrosis occurs in the IC in the acute phase, causing irreversible tissue loss while apoptosis occurs in the penumbra or peri-infarct region (PIR), which is a region between the lethally damaged IC and the normal brain. The PIR has partially preserved energy metabolism (moderately hypo-perfused) and retains structural integrity but has lost or impaired function. While cells in the IC can undergo permanent anoxic depolarization, cells in the PIR can repolarize with more energy consumption. This region can progress to infarction due to persistent excitotoxicity, spreading depolarization, inflammation, and apoptosis (Dirnagl et al. 1999). Therefore, the goal of neuroprotection is to salvage the ischemic penumbra. Alternative blood flow pathways, called collaterals, can sustain viability in penumbral regions, although the extent of collateral flow varies between individuals (Dirnagl et al. 1999). Patients with good collateral blood flow have infarcts that progress slower, while those with poor collateral blood flow display a more rapid progression of infarction.

Anoxic depolarization develops minutes after ischemia in neurons. As a result of the loss of membrane potential, voltage-dependent Ca²⁺ channels are activated, and excitatory amino acids, including glutamate, are released into the extracellular space (Campbell et al. 2019; Lo et al. 2003). Since energy-dependent reuptake is impeded, glutamate accumulation in the extracellular space is exacerbated. N-methyl-D-aspartate receptors (NMDARs) are blocked by extracellular Mg²⁺ under normal conditions; however, upon depolarization, the Mg²⁺ is removed, which leads to substantially higher conduction (Nowak et al. 1984) and subsequent Ca²⁺ influx and release from intracellular stores (Soriano et al. 2008; Wu and Tymianski 2018; Mayer and Miller 1990). Glutamate accumulation and subsequent Ca²⁺ overloading have diverse consequences. Ca²⁺ overloading in neurons following bioenergetic failure activates numerous Ca²⁺-dependent enzymatic reactions and subsequently induces lipolysis, proteolysis, breakdown of ion homeostasis, and disaggregation of microtubules, causing cell death and tissue damage in the IC during acute phase after ischemic stroke (Folbergrova et al. 1995). The Ca²⁺ increase also activates neuronal nitric oxide synthase (NOS) and results in free radical production and cell death processes such as apoptosis, necrosis, necroptosis, and autophagy. NO reacts with a superoxide anion to form peroxynitrite that promotes tissue damage due to its highly reactive nature (Iadecola et al. 1997; Barber and Demchuk 2003). Additionally, the activation of phospholipase A₂ and cyclooxygenase generates free-radical species leading to lipid peroxidation and membrane damage. An increase in free radicals leads to a leaky mitochondrial membrane and subsequent cytochrome C release, which triggers apoptosis. Studies have shown that mice whose iNOS gene expression was induced with mRNA had larger infarcts and motor deficits produced by occlusion compared to iNOS knockout mice (Iadecola et al. 1997).

Ca²⁺ increase was also observed in astrocytes in the acute phase following FIS. It was reported that astrocytes exhibited enhanced Ca²⁺ signaling in IC and PIR after photothrombosis (Ding et al. 2009), which may in turn induce more glutamate release from astrocytes. Deletion of IP₃R2 in astrocytes could reduce brain infarction and limit motor function deficits (Li et al. 2015; Dong et al. 2013). These results suggest that, in addition to neuronal Ca²⁺ signaling, astrocytic Ca²⁺ increase also contributes to brain damage through a non-cell autonomous effect.

There are two major modes of ischemic cell death: necrotic and apoptotic. Necrosis only occurs after exogenous insults. Following acute, permanent vascular occlusion, necrosis is the predominant form of cell death taking place in the IC, where the cells shrink and become very electron-dense (Lipton 1999). A hallmark of necrosis is the double-stranded breakdown of DNA into nucleosomal segments, which manifests as DNA laddering with fragments that are multiples of around 200 bp (Bonfoco 1995; Orrenius 1995). Overactivation of poly(ADP-ribose) polymerase (PARP), an NAD⁺-consuming enzyme, has been proposed to lead to necrosis by excessive energy loss (Eliasson et al. 1997; Yuan 2009). Deletion of PARP-1 or use of PARP-1 inhibitors is neuroprotective in vitro and in vivo following ischemia (Eliasson et al. 1997; Zhang et al. 1994). Calpain is a cytosolic Ca²⁺-activated protease and is known to be highly upregulated after ischemia (Vosler et al. 2011; Bano et al. 2005). Its activation from Ca²⁺ overloading under ischemic conditions contributes to neuronal cell death by cleaving multiple substrates such as cytoskeletal and associated proteins, kinases, phosphatases, membrane receptors, and transporters. Group I mGluR inhibitors 2-methyl-6-(phenylethynyl)pyridine (MPEP) and LY36738 could inhibit chaplain activation and thus reduce brain infarction (Li et al. 2013a).

Different from the necrotic death in the IC, neurons in the PIR may undergo apoptosis several hours or days after the onset of ischemic stroke. The protein families of Bcl-2 and caspase play critical roles in the activation, signal transduction, and execution of apoptosis (Broughton et al. 2009; Yuan 2009). Among the identified caspases, caspases 1, 8, and 9 appear to have key roles in ischemia-related apoptosis (Broughton et al. 2009; Yuan 2009). Caspases are activated when cytochrome C is released from the mitochondria and activates an apoptosome complex in the presence of dATP (Broughton et al. 2009). Activated caspases are aspartate-specific cysteine proteases that cleave enzymes and modify proteins which affect homeostasis and repair. Caspase activity can be blocked by the administration of small peptides which bind covalently to the catalytic pocket and alkylate the cysteine at position 70. Caspase inhibitors have been shown to decrease the volume of dead tissue in ischemia as well as neurological deficits (Hara et al. 1997). When ischemia is mild, inhibitors are particularly effective since they can be coupled with MK801, an NMDA-receptor antagonist, or growth hormones like fibroblast growth factor. While NMDA-receptor antagonists are typically administered before or immediately after ischemia, caspase inhibitors may still reduce injury even when injected many hours later. TNF1α, a cytokine, is also involved in apoptosis. TNF1α can exacerbate injury by causing a rapid decrease in mitochondrial membrane potential and thus impair mitochondrial function and increase caspase 8 activity, resulting in the release of cytochrome c from the mitochondria (Doll et al. 2015). Apoptosis-inducing factor (AIF) translocation from mitochondria to nuclei is involved in caspase-independent apoptosis (Wang et al. 2016; Broughton et al. 2009). NAD⁺ repletion or neuronal overexpression of NAMPT, the rate-limiting enzyme in the NAD⁺ salvage pathway, can reduce glutamate- and OGD-induced apoptosis through suppressing AIF translocation (Wang et al. 2014b, 2016). It is worth mentioning that one insult may lead to more than one mode of death in the same cell population, and cells may also manifest signs of multiple forms of death (Wang et al. 2016).

Astrocytes and Reactive Astrogliosis After Ischemic Stroke

Astrocytes are starlike cells and the most abundant glial cell type in the CNS. Classically, based on morphology and specific protein markers, there are two major types of astrocytes in the adult brain: fibrous astrocytes, present in white matter tracts such as the corpus callosum, and protoplasmic astrocytes, present in gray matter such as the cortex. Mature astrocytes have the following eight criteria (Kimelberg 2010): (a) electrically non-excitable; (b) a very negative membrane potential determined by the transmembrane K⁺ gradient; (c) expression of functional transporters for glutamate and GABA uptake; (d) a large number of intermediate filament bundles, which are the sites of the astrocyte-specific protein glial fibrillary acidic protein (GFAP); (e) having glycogen granules; (f) processes from each cell surrounding blood vessels; (g) many more processes from each cell surrounding synapses; and (h) linkage to other astrocytes by gap junctions consisting of connexins (CX) 43 and 30.

Astrocytes are traditionally considered as supporting cells for maintaining ionic homeostasis and providing growth factors for neurons and structural support in the CNS. Protoplasmic astrocytes exhibit the following functions under normal conditions (Kimelberg and Nedergaard 2010; Kimelberg 2010): (a) extracellular K⁺ buffering, (b) control of extracellular H⁺ and brain pH, (c) uptake of glutamate and GABA with their transporters, (d) mobilizing intracellular Ca²⁺ stores by activation of G-protein-coupled receptors (GPCRs) such as mGLuR5 and P2Y, (e) regulation of cerebral blood flow, (f) control of water transport by aquaporin (AQP) water channel, (g) astrocyte-neuron lactate shuttle, and (h) modulation and control of synaptic activity.

Although many astrocytes conform to the aforementioned criteria and functional roles, they are heterogeneous in morphology, molecular expression, and physiological function (Zhang and Barres 2010; Matyash and Kettenmann 2010). Morphologically, a protoplasmic astrocyte is highly branched and has several primary processes each with elaborated sub-branched fine process arborizations to form a bush-like territory with little overlap between neighboring astrocytes (Wilhelmsson et al. 2006; Bushong et al. 2002). A fibrous astrocyte has thicker and less branched processes with a high degree of overlap with neighboring astrocytes. Intermediate filament protein GFAP is primarily expressed in the thick main processes in astrocytes and has been considered as a “pan-astrocyte” marker. Transcriptomic study has revealed that the Aldh1L1 gene is the most widely and homogeneously expressed in astrocytes and that the Aldh1L1 protein is highly expressed in the cell body and extensive processes of an astrocyte (Cahoy et al. 2008). Therefore, Aldh1L1 is now considered as a new “pan-astrocyte” marker. Growing evidence indicates that astrocytes also play an active role through the “tripartite synapse,” which includes the pre- and post-synaptic neuron as well as the surrounding astrocyte. Astrocytes can modulate synaptic function, wakefulness, cognition, and memory (Araque et al. 1999; Halassa et al. 2007, 2009; Haydon 2001; Volterra and Meldolesi 2005; Khakh and Sofroniew 2015; Santello et al. 2019; Cui et al. 2018; Suzuki et al. 2011). Therefore, astrocytes are global controllers in the CNS which coordinate both local responses and those over larger distances.

Astrocytes are also involved in neurological disorders, including ischemic stroke (Seifert et al. 2006; Nedergaard and Dirnagl 2005; Maragakis and Rothstein 2006; Ding et al. 2009; Choudhury and Ding 2016; Ding 2014; Li et al. 2015). Ischemia induces a variety of molecular and cellular changes in astrocytes at the PIR, including cellular proliferation, morphology, and gene expression in a temporally and spatially dependent manner. Reactive astrogliosis and subsequent formation of a glial scar are the hallmarks of focal ischemic stroke (Lo 2008; Choudhury and Ding 2016; Ding 2014). The activated astrocytes are therefore called reactive astrocytes (RAs) that can be detected by enhanced expression of GFAP and many other proteins, as well as dramatic morphological changes (Choudhury and Ding 2016; Li et al. 2014; Sofroniew and Vinters 2010).

RAs in the PIR exhibit spatiotemporally dependent changes in morphology (hypotrophy), proliferation capacity, function, and gene expression during the subacute phase (Barreto et al. 2011; Li et al. 2014; Choudhury and Ding 2016). Morphologically, RAs become hypertrophic with large processes that can be revealed by GFAP staining. After a prolonged time following focal ischemic stroke, the morphology of RAs remains stable, the proliferation of RAs ceases, and a glial scar is formed (Barreto et al. 2011; Li et al. 2014; Ding 2014; Choudhury and Ding 2016; Burda and Sofroniew 2014; Voskuhl et al. 2009). Reactive astrogliosis and glial scar formation eventually cause substantial tissue remodeling and permanent structural changes in the penumbra. Analysis of immunostaining for GFAP and individually dyed cells in astrogliosis induced in the cortex or hippocampus by electrically induced lesion revealed that astrocyte processes become thicker and bushier (Wilhelmsson et al. 2006; Li et al. 2014; Zhang et al. 2020b).

The developing scar contains extracellular matrix components, and this extracellular proteoglycan deposition is part of the initial response that likely limits the spread of damage. The scar narrows as it matures, but persisting features remain, such as the upregulation of GFAP in astrocytes and their tightly intertwined processes. Selective impairment of scar formation leads to a greater spread of tissue damage and worse neurological outcomes. The impaired restoration of the blood-brain barrier leads to greater infiltration of leukocytes, inflammatory cell spread, and increased neuronal loss, which were observed when features of glial scar maturation were disrupted by transcription 3 (STAT3) in astrocytes (Wanner et al. 2013). Treatment with DAPT, which inhibits the Notch-activating enzyme γ -secretase, reduces damage to the ischemic brain, possibly by causing a reduction in the proliferation of reactive astrocytes. From this treatment or conditional knockout of the receptor, mice with impaired Notch signaling exhibited more microglia and macrophage invasion (Shimada et al. 2011; LeComte et al. 2015). It has been suggested that proliferation and differentiation of reactive astrocytes is promoted by Notch signaling via the translocation of the transcription factor Olig2 from the cytosol to the nucleus (LeComte et al. 2015).

On the other hand, the proliferation rate reaches a peak 3–4 days after FIS and recedes thereafter (Li et al. 2014; Barreto et al. 2011). Thus, metabolically, RAs are highly active on day 3–4 post-stroke to meet high energetic and biosynthetic demands for proliferation. Proliferation of RAs occurs mostly within 200μm from the IC and allows for the development of a glial scar after ischemia (Choudhury and Ding 2016; Zhang et al. 2020b; Li et al. 2014). Some astrocytes in the tissue adjacent to the infarct are actually derived from neural stem cells which migrate from the subventricular zone, and some survive as a part of the mature glial scar weeks after stroke (Faiz et al. 2015). Other cells, including microglia and/or macrophages, peri-vascular pericytes, or stromal cells, also proliferate (Barreto et al. 2011; Li et al. 2014; Fernandez-Klett et al. 2012). Many signaling pathways, including mitogen-activated protein kinases (MAPK), Notch signaling pathway, STAT3, and transforming growth factor beta (TGF-β) signaling, may participate reactive astrogliosis after ischemic stroke (Choudhury and Ding 2016).

Most changes in gene expression in RAs are due to altered cellular responses in the PIR, based on gene array analysis of RAs acutely isolated from brain tissue following MCA occlusion (Zamanian et al. 2012). For many genes, increased expression peaked 1 to 3 days after the onset of stroke, while a small proportion of genes including GFAP and some chemokines had increasing expression up to 1 week after stroke. Evidence also suggests that astroglia are influenced by specific conditions associated with the insult rather than an all-or-none response (Zamanian et al. 2012).

RAs have been demonstrated to have protective effects after FIS and injury (Linnerbauer and Rothhammer 2020; Myer et al. 2006; Faulkner et al. 2004; Anderson et al. 2016; Choudhury and Ding 2016). Proliferating RAs may impact tissue preservation, repair/remodeling, and functional outcome. Recent studies indicate RAs are also involved in depression, anxiety, cognition, and post-stroke mood shifting.

Post-stroke Symptoms and Post-stroke Mood Disorder (PSMD)

Mood disturbances are also frequent symptoms in stroke survivors (Kim 2016). After a stroke, many patients not only have some physical disability including difficulties in moving, speaking, and seeing, but it is common that patients may also exhibit changes in mood or emotion even after rehabilitation therapy (Schottke and Giabbiconi 2015). Anger, frustration, lack of motivation, and crying or laughing for the wrong reasons are also common for stroke patients. Post-stroke depression (PSD) (Robinson and Jorge 2015), post-stroke anxiety (PSA) (Maaijwee et al. 2016), and pseudobulbar affect (PBA) (Gillespie et al. 2016) are common post-stroke mood disorders (PSMD).

Symptoms of PSD include depressed mood, anhedonia, loss of energy, decreased concentration, and psychic retardation. It is characterized by feelings of overarching sadness, lack of pleasure, or changes in eating and sleeping patterns. Multiple studies have found that PSD affects between one-third to two-thirds of stroke survivors (Schottke and Giabbiconi 2015; Fang et al. 2017; Ayerbe et al. 2013; Kim 2016; Robinson and Jorge 2015; Virani et al. 2020), although its prevalence decreases over time. A systematic review of patients found that depression is associated with increased disability and mortality (Ayerbe et al. 2013).

PSA affects about 20% of survivors and occurs when they focus on worries and feel anxious for no particular reason (Schottke and Giabbiconi 2015; Wright et al. 2017). The core symptoms of PSA are excessive anxiousness or worry and difficulty in controlling worries, restlessness, decreased energy, poor concentration, irritation, nervous tension, and insomnia (Maaijwee et al. 2016). Meta-analyses have suggested that there is a significant correlation between PSD and PSA (Schottke and Giabbiconi 2015; Wright et al. 2017).

PBA is characterized by a mismatch between feelings and expression. Individuals may cry or laugh in an uncontrolled manner. Crying is a more common PBA presentation following stroke than laughing (Gillespie et al. 2016). PBA affects approximately one in five stroke survivors at the acute and post-acute phases and one in eighth survivors 6 months or more post-stroke. Research indicates that PBA is more common in survivors of brainstem stroke, but it can also occur with other types of strokes (Balakrishnan and Rosen 2008).

Tools for cognitive and mood assessment have been suggested (Quinn et al. 2018). A framework for assessment emphasizes the need for differing approaches to testing at differing points in the stroke pathway rather than a comprehensive critique of all cognitive and mood assessment tools. Cognitive and mood problems are both associated with poor outcomes of stroke. Clinically, a number of drug and psychosocial treatments have been assessed, but the results have been disappointing. These drug trials are generally of poor quality and do not provide sufficient information to judge their true costs and benefits (Hackett et al. 2008). Psychosocial interventions are popular with patients, but there is conflicting evidence for their effectiveness in either treating or preventing anxiety and depression (Gao et al. 2016; Wu et al. 2012). The problem-based and behavioral therapies seem to be promising for patients with stroke (Hill et al. 2019). Improved coping skills should result in reduced psychological distress and rates of depression. A licensed mental health practitioner (therapist or nurse) and the individual actively worked together toward recovery based on a psychological assessment. Standardized measures of mood (28-item General Health Questionnaire/GHQ-28), cognitive state (mini-mental state examination), and function (Barthel ADL Index, Frenchay Activities Index) were taken at different times. Hill et al. reported that 6 months later, all psychological and activity measures favored problem-solving therapy. At 12 months, patients in the problem-solving therapy group had significantly lower GHQ-28 scores and lower median Present State Examination symptom scores (Hill et al. 2019). In addition, the problem-solving therapy group was more satisfied with some aspects of care.

Astrocytes in Major Depressive Disorder (MDD) and PSMD

MDD is the most prevalent form of depression, often manifested with a long-lasting and recurrent psychiatric condition. It is reported that MDD affects 20% of the population throughout their lifetimes (Kessler and Bromet 2013), and it is now considered the leading cause of disability worldwide. The symptoms of MDD included depressed (low) mood; anhedonia; feelings of hopelessness (despair), worthlessness, or guilt; changes in appetite, weight, and sleep; an inability to feel pleasure (anhedonia); fatigue; and suicidal ideation. Studies suggest that the neural activities of specific brain circuits are altered in response to external stimuli, such as stress, as a result of maladaptive molecular and cellular changes in MDD. Astrocytes interact intimately with neurons to support and regulate essential functions and mediate information processing through tripartite synapses in the brain, where astrocyte processes wrap tightly around pre-synaptic and post-synaptic sites (Araque et al. 1999; Halassa et al. 2007; Haydon 2001; Volterra and Meldolesi 2005; Khakh and Sofroniew 2015). Therefore, astrocytes have been receiving increasing attention in mood disorders since significant abnormalities were observed in the postmortem brain of MDD patients. Indeed, studies using postmortem brain specimen and animal models of depression provide a large body of evidence that astrocytes play an important role in MDD and PSMD.

Astrocyte in MDD

Studies from Human Postmortem Brain Specimens

Brain imaging studies have revealed marked volume reductions in the hippocampus and medial prefrontal cortex (mPFC) in MDD (Drevets 2000; Sheline 2003). Studies from human specimens showed that abnormalities in glial cells may alter normal brain function and likely contribute to mood disorder development. Profound alterations of astrocytes connecting to mood disorders were observed from postmortem brain specimen. These include changes in cell number and cell morphology in patients who had mood disorders. Prominent reductions in glial cell number and packing density have been reported in independent laboratories based on studies using postmortem brains from subjects with mood disorders in different regions of the PFC, the anterior cingulate cortex (Gittins and Harrison 2011; Cotter et al. 2001a), and the amygdala (Rajkowska 2000; Altshuler et al. 2010; Ongur et al. 1998; Bowley et al. 2002). Such a striking cellular deficit in glial number suggests that glia may be unique targets for novel strategies in the treatment of mood disorders.

Altered expression of astrocyte-specific biomarkers was also observed in postmortem brain specimens of individuals that had suffered from mood disorders, for example, low levels of GFAP have been found in the hippocampus, PFC, anterior cingulate, and amygdala (Webster et al. 2001; Gittins and Harrison 2011; Altshuler et al. 2010). In postmortem brains of suicide completers, reduced GFAP mRNA and protein in the mediodorsal thalamus and caudate nucleus were observed in depression-related suicides compared with controls, suicides not linked to depression (Torres-Platas et al. 2016). Furthermore, a regional comparison revealed that GFAP expression in both subcortical regions was, on average, between 11- and 15-fold greater than in the cerebellum and neocortex. Examining astrocyte morphology by immunohistochemistry showed that astrocytes in both thalamus and caudate displayed larger cell bodies and extended more ramified processes across larger domains than the previously described cortical astrocytes. This study reveals that astrocytic abnormalities are not brain wide and suggests that they are restricted to cortical and subcortical networks known to be affected in mood disorders.

S100B proteins are a family of acidic proteins that influence cellular responses along calcium signal transduction pathways primarily produced by astrocytes. It was found that S100B was elevated in the blood serum and cerebrospinal fluid of MDD patients, and antidepressant treatment could lower S100B levels in parallel with reduced depressive symptoms (Matthias et al. 2013; Schroeter et al. 2002). In addition, a systematic and quantitative meta-analysis demonstrated that both young and older subjects suffering from mood disorders showed elevated S100B values in blood serum compared to control subjects. Also, higher levels of S100B were found in older compared with younger adult subjects, indicating that glial pathology is modified by age in mood disorders (Schroeter et al. 2011). Thus, S100B can be considered as a new marker for MDD.

Changes to gene transcription and protein expressions relating to normal astrocyte function were also found in patients diagnosed with mood disorders. For example, gene and protein expression of some astrocytic function-related proteins, including glutamine synthetase, glutamate transporters, and even gap junction proteins, was downregulated in patients with depression (Sequeira et al. 2009; Bernard et al. 2011). Altered cortical glutamatergic and GABAergic signal transmission in depression is associated with downregulation of high-affinity glutamate transporters GLT1 and glutamine synthetase (Choudary et al. 2005). Using immunostaining, a reduction of blood vessel coverage by AQP4 astrocytic endfeet was reported in MDD subjects (Rajkowska et al. 2013).

Postmortem histopathologic studies of depressed patients strongly suggest that the abnormalities in astrocytic functions, such as regulation of water homeostasis, blood flow, glucose transport and metabolism, the blood-brain barrier, glutamate and GABA turnover, and synaptic plasticity, may contribute to the pathophysiology of MDD.

Studies from Animal Models

While studies using postmortem brain specimens provide insights into pathological changes of astrocytes in MDD, studies using animal models may provide the underlying molecular mechanisms by which astrocytes participate in disease etiology. Stressful life events including acute and chronic stress increase the risk for MDD in humans (Arsenault-Lapierre et al. 2004). Thus, in animal experimental studies on MDD, mice and rats are exposed to acute stress (e.g., tail suspension test, forced swim test, unavoidable foot shock test, social defeat test, or restraint stress) or chronic unpredictable stress (CUS) to induce depressive-like states (Henn and Vollmayr 2005).

Due to the nature of tripartite synapses, astrocyte-derived substances such as glutamate, D-serine, lactate, and ATP can actively modulate synaptic function. Meanwhile, the changes in the expression of astrocyte-specific glutamate transporters, such as GLT1 and GLAST1; glutamate receptors, such as mGluR5; enzymes, such as glutamate synthesis and lactate dehydrogenase; and ion channels and gap junctions may affect/regulate ion homeostasis, synaptic transmission, pH, redox state, etc. These multiscale and multimodal interactions of astrocytes with synaptic circuits are likely to contribute to behavior. Because neuronal synaptic changes are a hallmark of depression and astrocytes are integral to tripartite synapses, astrocytes are very likely to participate in the pathology of depression and anxiety disorders. Indeed, a large number of studies from rodent model have provided evidence of astrocytic involvement in MDD. For extensive reviews of astrocytes in this area, readers are advised to consult a few detailed reviews (Wang et al. 2017; Bender et al. 2016; Park and Lee 2020). Here, I will briefly summarize the recent progress in this area.

An attractive hypothesis for MDD is that impaired ATP release from astrocytes causes MDD (Illes et al. 2020; Cao et al. 2013). Cao et al. identified ATP as a key factor involved in astrocytic modulation of depressive-like behavior in the hippocampus and prefrontal cortex of adult mice with chronic social defeat stress (Cao et al. 2013). Low ATP abundance was observed in the brains of mice that were susceptible to chronic social defeat. Furthermore, they found that the administration of ATP induced a rapid antidepressant-like effect in these mice. Both a lack of inositol 1,4,5-trisphosphate receptor type 2 (IP₃R2) and transgenic blockage of vesicular gliotransmission-induced deficiencies in astrocytic ATP release caused depressive-like behaviors that could be rescued via the administration of ATP. Using transgenic mice that express a Gq G-protein-coupled receptor (GPCR) only in astrocytes to enable selective activation of astrocytic Ca²⁺ signaling, they also found that stimulating endogenous ATP release from astrocytes induced antidepressant-like effects in mouse models of depression. Moreover, P2X2 receptors in the medial prefrontal cortex mediated the antidepressant-like effects of ATP. Their results highlight astrocytic ATP release as a biological mechanism of MDD. However, the data above using IP₃R2-deficient mice is inconsistent with a study by Petravicz et al. (Petravicz et al. 2014), who found that the mice did not show any abnormalities in the open field test or the tail suspension test. Therefore, it remains difficult to clarify the relationship between astrocytic Ca²⁺ signaling and depression, suggesting a complex nature of depression and the lack of decisive behavior tests to evaluate it.

The lateral habenula (LHb) is a nucleus that relays information from the limbic forebrain to multiple monoamine centers and has recently emerged as a key brain region in regulating negatively motivated behavior and the pathophysiology of major depression (Cui et al. 2018; Li et al. 2013b; Hu et al. 2020; Baker et al. 2016; Yang et al. 2018). In a rat depression model of congenitally learned helplessness, Cui et al. recently discovered that Kir4.1 in astrocytic membrane processes was upregulated in the lateral habenula (LHb) (Cui et al. 2018). Their study using electrophysiology and modeling data shows that the level of Kir4.1 in astrocytes tightly regulates the degree of membrane hyperpolarization and the amount of bursting activity of LHb neurons. Astrocyte-specific gain and loss of Kir4.1 in the LHb bidirectionally regulates neuronal bursting and depression-like symptoms. Together, their results show that a glia-neuron interaction at the perisomatic space of LHb is involved in setting the neuronal firing mode in models of a major psychiatric disease. Therefore, targeting Kir4.1 in the LHb might be a potential strategy for treating clinical depression.

Neuroinflammation contributes to the cognitive impairments accompanying many neurological disorders. Chronic stress and neuroinflammation are considered to be fundamental in the etiology of MDD. Astrocyte dysfunction and inflammation have been proven to be associated with the pathogenesis of MDD. Using mouse depression models of 6 weeks of chronic unpredictable mild stress (CUMS) or 10 days of lipopolysaccharide (LPS) intraperitoneal injection, Leng et al. explored intermediary components that are modulated by stress and neuroinflammation (Leng et al. 2018). They found that multiple endocrine neoplasia type 1 (Men1; protein: menin) expression is attenuated in the brain of mice exposed to CUMS or LPS. Astrocyte-specific deletion of Men1 (GcKO) led to depressive-like behaviors in mice and enhanced IL-1β production through NF-kB activation. Further, they observed that depressive-like behaviors in GcKO mice could be restored by an NF-kB inhibitor or an IL-1β receptor antagonist. Importantly, they identified a SNP in human MEN1, where G503D substitution is associated with a higher risk of MDD onset. G503D substitution abolished menin-p65 interactions, thereby enhancing NF-kB activation and IL-1β production. The study revealed a distinct role of tumor suppressor menin in astrocytes in regulating astrocytic inflammation in depression, and menin may be an attractive therapeutic target in MDD.

IL-10 is a key cytokine that is mainly produced by astrocytes and microglia and that represses excessive inflammatory responses. In the CNS, IL-10 is upregulated after various insults. Astrocytes isolated from transgenic mice deficient with IL-10 are prone to characteristic A1 reactive astrocytes and these transgenic mice exhibited increased immobility time in the forced swim test and defective learning and memory behavior in the Morris water maze test (Zhang et al. 2020a), suggesting IL-10 contributes to the depression-like behavior and memory deficits.

It was found that local application of TFNα at pathological levels activates astrocyte TNF receptor type 1 (TNFR1), which in turn triggers an astrocyte-neuron signaling cascade that results in persistent functional modification of hippocampal excitatory synapses. Astrocytic TNFR1 signaling induced hippocampal synaptic alteration and contextual learning-memory impairment. This process may contribute to the pathogenesis of cognitive disturbances (Habbas et al. 2015). High extracellular levels of TNFα can trigger release of glutamate from astrocytes (Santello et al. 2011). Thus, the mechanism by which the cytokine TNFα affects cognitive disturbances upon CNS inflammation is via a local increase of TNFα in the hippocampal dentate gyrus that activates TNFR1, which then triggers an astrocyte-neuron signaling cascade resulting in a persisting modification of hippocampal excitatory synapses (Habbas et al. 2015). Astrocytes are not only key to normal cognitive functions, but also the modification of synapses following neuroinflammation.

Astrocytes provide metabolic support to neurons. One mechanism is through the astrocyte-neuron lactate shuttle (Pellerin and Magistretti 1994; Magistretti and Pellerin 1996; Belanger et al. 2011; Magistretti and Allaman 2015). It is reported that peripheral administration of lactate exerts antidepressant-like behavioral effects (Carrard et al. 2018). These behavioral effects of lactate may be brought about by its ability to increase synaptic excitability.

The glutamatergic predominance in the excitatory-inhibitory balance is postulated to be involved in the pathogenesis of depression. Such an imbalance may be induced by astrocyte ablation which reduces glutamate uptake and increases glutamate levels in the synaptic cleft, causing depression-like behaviors. A causal relation between astrocytic dysfunction and depression is also provided by animal studies. The selective destruction of frontocortical astrocytes with the gliotoxin L-α-aminoadipic acid (L-AA) is sufficient to trigger a depressive-like phenotype (Banasr and Duman 2008; Domin et al. 2014). Both the behavioral and GFAP level changes could be prevented by injection of 3-((2-methyl-4-thiazolyl)ethynyl)pyridine (MTEP), a mGluR5 antagonist, through the inhibition of glutamatergic transmission (Domin et al. 2014). The results demonstrate that glial ablation in the PFC is sufficient to induce depressive-like behaviors similar to chronic stress, supporting the hypothesis that loss of glia contributes to the core symptoms of depression. Astrocyte dysfunction therefore may lead to excitatory-inhibitory imbalances, resulting in mood disorders such as depression and anxiety. Astroglial degeneration in the prefrontal cortex is a useful rat depression model.

Gap junctional communication is a main determinant of astrocytic function. Rats exposed to CUS, a rodent model of depression, showed behavioral deficits in sucrose preference test (SPT) and novelty suppressed feeding test (NSFT) and exhibited significant decreases in diffusion of gap junction channel-permeable dye and expression of Cx43, a major component of astrocyte gap junctions, and abnormal gap junction ultrastructure in the PFC (Sun et al. 2012). The results showed that infusions of gap junction blocker carbenoxolone (CBX) induced anhedonia in SPT and anxiety in NSFT, a core symptom of depression. Gap junction dysfunction contributes to the pathophysiology of depression.

Water channel AQP4 is expressed in astrocytes in the brain and is also involved in the pathogenesis of depression. Using a mouse model of depression induced by repeated corticosterone injections (Zhao et al. 2008), AQP4 knockout mice exhibit exacerbated depression-like behaviors based on forced swimming test (FST) and tail suspension test (TST) (Kong et al. 2014). These knockout mice also show a significant loss of astrocytes; aggravated downregulation of excitatory amino acid transporter 2 (EAAT2), synapsin-1, and glial cell line-derived neurotrophic factor (GDNF); and less hippocampal neurogenesis. Thus, astrocytic AQP4 can modulate astrocytic function and adult neurogenesis during the pathogenesis of depression and might be a potential target for the treatment for depression.

Blockade of astrocytic glutamate transporter GLT1 induces depressive-like behaviors in rats (Bechtholt-Gompf et al. 2010; John et al. 2012). Dysfunction or imbalance of the monoamine- or L-glutamate (L-Glu)-mediated synaptic transmission is known to be a pathogenic cause of many disorders. Astrocytes take up synaptic L-Glu during synaptic transmission through excitatory amino acid transporters GLT1 and GLAST. L-Glu is subsequently metabolized to L-glutamine (L-Gln) by glutamine synthetase (GS). Then, release of astrocytic L-Gln is used as a neuronal L-Glu precursor, making up the glutamine cycle. Expression levels of astrocyte transporters and metabolizing enzymes are dynamically regulated by synaptic activity, which empowers astrocytes to support synaptic transmission. Pharmacological blocking of astrocyte glutamate buffering by blocking GLT1 triggered depressive behaviors in the form of latency to drink sucrose solution, increased intracranial self-stimulation, or decreased social interaction (John et al. 2012). Decreased GLT1 and GLAST expression may cause impaired L-Glu turnover, which contributes to depression. Evidence that riluzole, which activates L-Glu transporters, reverses the decreased GFAP expression in rat prefrontal cortex and improves depressive-like behavior supports the involvement of impaired L-Glu turnover in depression pathogenesis (Banasr and Duman 2008). The role of astrocytes in the excitatory-inhibitory imbalance hypothesis of depression and anxiety is related to changes in glutamate receptor function.

Depression and anxiety are associated with NMDAR, and its antagonist ketamine was shown to produce a rapid antidepressive effect (Yang et al. 2018; Li et al. 2010). NMDARs are activated after binding of the agonist glutamate to the NR2 subunit along with a co-agonist, either L-glycine or D-serine, to the NR1 subunit at a glycine modulatory site to function as a gatekeeper. L-serine is synthesized by astrocytes, which is then transported to neurons for conversion to D-serine by serine racemase (SR). Substantial evidence suggests that D-serine is the most relevant co-agonist in forebrain regions, supporting its role in fear conditioning and anxiety disorders (Wolosker and Balu 2020). Transgenic mice overexpressing SR which increase the availability of astrocyte D-serine led to a reduced depressive phenotype based on the forced swim, novelty suppression of feeding and olfactory bulbectomy paradigms; chronic dietary D-serine supplement mimics the depression-related behavioral phenotype observed in SR transgenic mice (Otte et al. 2013). The acquisition and extinction of fear memory engages the SR/D-serine system in the mouse amygdala, and D-serine administration facilitates fear extinction (Balu et al. 2018).

Overall, the aforementioned studies indicate that astrocytes play a role in depression-like behaviors using different mechanisms, and impaired astrocytic function is necessary and sufficient to induce depression-like behaviors in animal experimental studies.

Astrocytes in PSMD

Although there were few functional studies on the role and mechanism depression in humans, studies from human specimens and animal models of MDD may provide insights into the mechanism and therapeutic strategies for PSMD. Generally, there are two causes of mood swings after stroke: biological changes and lifestyle changes. Biological changes are caused by damage in the emotion centers of the brain. Few studies have been done on the relationship between biological changes and PSMSD at cellular and molecular levels. Data from experimental studies using rodent models might provide insight on PSMDs, depression, and anxiety in general. Since MRI studies have found significant alterations in different brain regions in MDD patients, such as in the frontal lobe, hippocampus, temporal lobe, thalamus, striatum, and amygdala (Pandya et al. 2012; Zhang et al. 2018), whether stroke patients develop PSMD might be dependent on the region that is injured. On the other hand, ischemic stroke causes changes in morphology, metabolism, and gene expression in reactive astrocytes. Any changes related to glutamate synthesis, transportation, and degradation after stroke may affect mood swings.

The monoaminergic hypothesis of depression postulates symptoms to be a result of an imbalance in the central monoaminergic system including serotonergic, dopaminergic, and/or noradrenergic neurotransmission (Marathe et al. 2018). Astrocytes express transporters for norepinephrine and serotonin (Inazu et al. 2003; Hirst et al. 1998), which are the targets of several classical antidepressant drugs. This raises a possibility that antidepressants can have direct effects on astrocytes. These studies suggest that astrocytes may exert control of serotonergic and noradrenergic transmission and are thus cellular targets for antidepressant drugs that block the reuptake of monoamines by astrocytes.

Wang et al. developed rat model for post-stroke depression (PSD) using middle cerebral artery occlusion (MCAO), followed by an 18-day chronic mild stress (CMS), and assessed depression-like behavior and the effects of the antidepressant citalopram (Wang et al. 2008). Using the open-field test (OFT) and the sucrose consumption test, they found that citalopram could ameliorate the behavioral abnormalities, suggesting that the ischemic rat CMS model is an appropriate model for PSD.

Inflammation and alterations in glutamate neurotransmission are two novel pathways to pathophysiology in mood disorders (Haroon et al. 2017). Stroke causes dysfunction of glutamate transporters and reduced glutamate uptake by astrocyte in acute phase. Increasing data indicate that inflammation causes impaired astrocytic glutamate uptake, and patients with depression and anxiety have increased inflammation (Haroon et al. 2017). Since inflammation also occurs in ischemic stroke, inflammation-induced impairment of glutamate uptake may contribute to the development of or enhanced predisposition for PSMD. Elevated glucocorticoids due to illness-related stress could also downregulate glial activity and exacerbate or predispose patients to psychiatric illness via enhanced excitotoxicity (Cotter et al. 2001b). Since excitotoxicity is a common acute consequence of ischemic stroke, this may contribute to the subsequent development of mood disorders such as post-stroke depression and anxiety.

Chen et al. showed that post-stroke enrichment environment (EE) increased high-mobility group box-1 (HMGB1) and interleukin-6 (IL-6) expression in astrocytes, led to decreased depression and anxiety-like behavior, and promoted angiogenesis and functional recovery compared to standard environment (Chen et al. 2017). EE mice treated with glycyrrhizin decreased, whereas EE mice treated with recombinant HMGB1 (rHMGB1) increased in the levels of IL-6 and p-AKT. Their study highlighted the role of the astrocytic HMGB1-IL6 pathway in PSMD in animal models.

Yu et al. found that GLT1 was downregulated in astrocytes in the post-stroke rat model (Yu et al. 2019b). Reduced expression of GLT1 in PSD astrocytes inhibited the formation of functional synapses by influencing glutamate metabolism. Another study from the same group investigated the effect of ceftriaxone, which increases GLT1 expression, on depression-like behaviors of rats after MCAO (Yu et al. 2019a). They found that treatment of ceftriaxone gradually increased GLT1 both in transcription and translation levels and inhibited depression-like behaviors by increasing locomotor and rearing activity and improving anhedonia of the rats. Moreover, ceftriaxone can promote glutamate circulation and synaptic plasticity by increasing astrocytic GLT1 levels. Their study indicates that reduction of GLT1 in astrocytes is one potential mechanism of pathogenesis of post-stroke depression.

Rats injected with the astrocyte-specific toxin L-AA in the mPFC, which is anatomically and functionally linked with cognitive and emotional processing, resulted in a pronounced loss of astrocytes in the region and adversely affected set-shifting, working memory, and reversal learning functions (Lima et al. 2014). The lesion sites also showed progressive neuronal loss and dendritic atrophy in surviving neurons, suggesting that L-AA-induced astrocytic loss in this brain region leads to neuronal damage that results in cognitive impairment. Therefore, it is clear that astrocytes play a key role in cognitive impairment, which also occurs after stroke.

The neurotrophic hypothesis in depression and anxiety disorders was proposed because lower serum levels of neurotrophic factors like brain-derived neurotrophic factor (BDNF) are often observed in patients with depression, while increased expression of BDNF and GDNF is observed in patients as a response to antidepressant treatment (Pisoni et al. 2018). BDNF overexpression in hippocampal astrocytes leads to antidepressant-like activity and increased neurogenesis in mice (Quesseveur et al. 2013). Recently, a study demonstrated that RAs expressed increased GDNF (Zhang et al. 2020b), and this suggests that RAs might play a role in improving PSMD through GDNF release.

D-Serine has also been found to play a role in post-traumatic stress disorder (PTSD) (Wolosker and Balu 2020). It was found that the single nucleotide polymorphism, rs4523957, in human serine racemase gene, is associated with PTSD based on postmortem human brain study (Balu et al. 2018). D-Serine might play a role in PSMD since stroke and traumatic injury have similar mechanisms of brain injury.

Overall, these data suggest that astrocytes/reactive astrocytes are an integral part of depression and anxiety through different mechanisms and may be a potential therapeutic target for PSMD.

Concluding Remarks

Ischemic stroke is a leading cause of human disability, but recovery of behavioral function in animal stroke models and young patients can be remarkable, much of which is due to neuroplasticity that involves the strengthening of existing synapses, synaptogenesis, or local sprouting. However, these effects are often accompanied by neurological problems because of failure to recruit larger and more diffuse networks to function normally. Beyond the disability in normal motor function, it is common that stroke survivors have depression and anxiety-like mood disorders. Although a large body of evidence indicates that astrocytes play a role in MDD from studies of both human postmortem specimen and animal models, there are fewer studies regarding the participation of astrocytes or reactive astrocytes in PSMD. Considering that astrocytes can influence neuronal function through a variety of mechanisms, future studies on the mechanisms by which astrocytes/reactive astrocytes play a role in PSMD should focus on the context of astrocyte-neuron interactions in stroke models.