Astrocytes, five times of neurons in number, play an important role in neuronal nutrition, support, protection and mutual signal transduction. On one hand, the protrusions of Ast surround the neurons and communicate with neurons and synapses. On the other hand, Ast wraps the blood vessels with the end feet and mediates the transmission of the biological information between neurons and blood vessels. Ast can not only regulate the transmission and strength of synapses, but also participate in the synaptic formation and neuronal differentiation (Stogsdill et al., 2017). Studies have shown that estrogen released by cultured primary Ast increased the number of newly formed synapses and enhanced synaptic signal transmission by activating the estrogen receptor-α on neurons (Fester and Rune, 2015). Thrombospondins-1/2 secreted by Ast promote the synaptic formation in the central nervous system (CNS) both in vivo and in vitro (Kim et al., 2017). Cholesterol released by Ast promotes synaptic formation and affects the synapse number during the development of neuron (Mauch et al., 2001). The most interesting is that rat embryonic stem cells could differentiate into neurons only in the medium containing Ast (Nakayama et al., 2003). In addition, Ast can provide energy and nutrients for neurons, and release citrate, an important substance supplying neuronal energy during hypoglycemia (Meshitsuka and Aremu, 2008) and produce lactic acid through glycolysis to provide some energy to neurons (Takahashi, 2020).

As the innate immune cells in the brain, MG monitor the brain microenvironment, serving as the first line of the CNS defense (Li and Barres, 2018). Generally, MG and Ast are activated into two states: the neurontoxin phenotype (M1/A1) and the neuroprotection phenotype (M2/A2), corresponding to either the destructive or reparative functions in the NVU, respectively (Liu et al., 2020). Activated MG induce A1 polarization of Ast by secreting interleukin (IL)-1α, tumor necrosis factor-α (TNF-α) and complement component 1q (C1q). These cytokines are necessary and sufficient to induce A1 Ast, which lose the ability to promote neuronal survival, outgrowth, synaptogenesis and phagocytosis, inducing the death of neurons and oligodendrocytes (Liddelow et al., 2017). Similarly, MG can also release inflammatory factors such as IL-1β to block the production of sonic hedgehog by Asc, but to drive the secretion of CXCL2 and CCL2, further disrupting the integrity of the BBB and increase neuroinflammatory responses. Ast can release a large amount of IL-17A after stroke, and at the same time, IL-17AR is also highly expressed, and the transformation of MG to M1 type can be reduced by inhibiting IL-17A derived from Ast, which suggests that the polarization of M1-type microglia may be mediated by the IL-17 signaling and is closely related to Ast (Ronaldson and Davis, 2020). MG and Ast can also interact in a paracrine manner by secreting cytokines such as transforming growth factor-β (TGF-β), IL-1, ATP, etc. (Liu et al., 2011). Moreover, a specific pyrimidine-receptor related pathway between MG and Ast, may affect chronic inflammation of the nerve and reaction of glia (Quintas et al., 2014). In addition, MG also affect Ast in the NVU upon stimulation by LPS, whereby MG are activated to produce NO, which subsequently induces neuroprotective effects in Ast through the Keap1/Nrf2 pathway (Takahashi, 2020). MG also release IL-10 to act on reactive astrocytes (RAs), which allow Ast to release cytokines and to play an active protective role in NVU (Zagrean et al., 2018).

As a complex composed of the brain endothelial cells, pericytes, extracellular matrix basal lamina and astrocytic end-feet (Najjar et al., 2013), BBB is an important structure in the NVU and a natural barrier for the brain to maintain the microvascular homeostasis. Ast, one of the three types of cells involved in the formation of BBB, stretch out to wrap more than 90% of capillary endothelial cells and pericytes, interact with brain microvascular endothelial cells and maintain the integrity of BBB (Thurgur and Pinteaux, 2018). One of main structures of BBB is the endothelial cells and the tight junctions (TJs) between Ast. Ast can express endothelial transport molecules such as glucose transporter-1, and its end-feet can release the signal molecules to support the formation and maintenance of TJs. Inside BBB, Ast are interconnected with neighboring cells through the channels of gap junction that are regulated by extra- and intra-cellular signals and allow exchange of information (Bell et al., 2020). Ast-endothelial gap junctions are mediated by connexin-43 (CX43) hemichannels that allow cell-cell transfer of nutrients, neurotransmitters and maintain water and ion balance inside and outside blood vessels. In this process, there are complex signal exchanges between Ast, pericytes and endothelial cells (Haj-Yasein et al., 2011; He et al., 2013; Nuriya et al., 2013). Prat et al. (2001) found that cultured endothelial cells alone cannot form BBB unless co-cultured with Ast or glial cell culture fluid. Willis et al. (2004) performed a rat model of Ast injury and found that Ast was injured and died at 4–24 h, while BBB permeability increased after 24 h, and then gradually returned to normal at 6–8 days after BBB injury. Therefore, it can be speculated that Ast play an important role in the regulation of BBB permeability. Moreover, interaction between Ast and endothelial progenitor cells (EPCs) may mediate neurovascular remodeling after stroke (Hayakawa et al., 2012).

Pericytes are very important components of the NVU serving as integrators, coordinators and effectors of neurovascular functions, including the maintenance of BBB integrity (Zheng et al., 2020), regulation of cerebral blood flow (CBF) (Kisler et al., 2017a), clearance and phagocytosis of cellular debris (Sá-Pereira et al., 2012). The contraction and relaxation of pericytes play a key role in the regulation of local microvascular blood flow. Some studies have shown that a proportion of pericytes are contractile, responsive to brain-generated vasoactive signals, and are able to influence capillary diameter both in vitro and in vivo (Hall et al., 2014; Mishra et al., 2016), which can affect the perfusion of local brain and the redistribution of blood flow (Sagare et al., 2013; Kisler et al., 2017b; Montagne et al., 2018). Using two-photon microscopy to observe and manipulate brain capillary pericytes in vivo, Hartmann et al. (2021) found that vasoconstriction is associated with a reduction in red blood cell velocity and flux in the same capillary, suggesting that induced contraction of pericytes can alter CBF. As the key cells that regulate the contraction and the relaxation of brain microvasculature, pericytes can affect the formation and permeability of BBB and participate in the reperfusion of blood flow in the brain, which may be, in this sense, closely related to the fact that Ast can affect the Ca²⁺ concentration (Iadecola, 2017). Armulik et al. (2010) found that pericyte deficiency increases the permeability of the BBB, indicating that pericytes play an important role by integrating endothelial cells and Ast, and regulation of the BBB in NVU.

Extracellular matrix, secreted by neurons or glial cells, surrounding the dynamic structure of cells, provides support for cell arrangement, and performs a series of physiological functions within NVU (De Luca et al., 2020). As one of the components of NVU, ECM can maintain the homeostasis through constant exchanges of water, irons and nutrients in quantity and quality, which are related to the activities of specific enzymes responsible for the degradation of ECM, such as matrix metalloproteinases (MMPs) (Bonnans et al., 2014), Zn²⁺ dependent ECM remodeling endopeptidases (Kucera et al., 2016), playing an important role in the degradation of ECM and the regulation of cell-cell adhesion, migration, infiltration, proliferation, apoptosis (Quattromani et al., 2018), and neuronal synapse remodeling (Oohashi et al., 2015; Stanic et al., 2016; Peng et al., 2018). Studies have confirmed that Ast overexpress MMP-2 and MMP-9 after ischemic stroke, which destroy the structure of ECM and increase the permeability of the BBB (Keep et al., 2014). The main function of MMP-2 is to degrade the basement membrane of the ECM, while the MMP-9 is to degrade and remodel the dynamic balance of the ECM. Min et al. (2017) found that heme can stimulate Ast to release TLR2, then increase the expression of MMP9, causing the damage of ECM and BBB. Some cytokines secreted by Ast can cause changes in the composition and structural integrity of the ECM (De Luca et al., 2018), and conversely, the destruction of the ECM can also significantly influence the function of Ast.

ERK1/2 is a key regulator in the MARK signal pathway and plays an important role in cell proliferation and differentiation. Studies have shown that activation of the ERK1/2 signal pathway also has dual effects to ischemic stroke (Maddahi and Edvinsson, 2010). On one hand, inflammatory factors or oxidative stress activates ERK1/2, which aggravates NVU damage; on the other hand, after the activation of ERK1/2 signal pathway, neurotrophic factors secreted by Ast play a neuroprotective role (Sawe et al., 2008). Some drugs targeting Ast can affect ischemic stroke through the ERK1/2 pathway. For example, Potassium2-(1-hydroxypentyl)-benzoate (D, L-PHPB), a new drug candidate for ischemic stroke at the phase II clinic trial. By up-regulating ERK activity, D, L-PHPB significantly increases the levels of BDNF and NGF secreted by Ast to promote neuronal survival and inhibit neuronal apoptosis under OGD/R conditions in the co-culture system of neurons and Ast (Liu et al., 2017). D, L-PHPB treatment suggests that the neuroprotective effects may be related to Ast (Zhang et al., 2006; Hu et al., 2012; Zhao et al., 2013; Peng et al., 2014), but the mechanisms of D, L-PHPB remain to be further studied. Tanshinone IIA inhibits the accumulation of hypoxia inducible factor-1α and stromal cell derived factor-1 in Ast under OGD conditions, the activation of ERK1/2 and AKT, and the proliferation of Ast (Huang X. et al., 2014), thereby reducing the degree of injury.

TLR4 is the earliest discovered Toll-like protein. After cerebral ischemia, TLR4 is firstly activated in both Ast and MG. NF-κB is one of the important downstream signaling nuclear transcription factors of TLR4. Some cytokines such as IL-2, IL-1β and TNF-α in the cell can activate the NF-κB in turn, thereby stimulating further cascade amplification of inflammatory signals. Inhibiting the activation of the TLR4/NF-κB signal pathway improves the damage of ischemic stroke (Zhao et al., 2018). Studies have shown that ginkgo diterpene lactones attenuates ischemic injury by inhibiting the TLR4/NF-κB pathway, inhibiting platelet aggregation, astrocyte activation and pro-inflammatory cytokine production (Li et al., 2020). Pien Tze Huang (PTH) significantly reduces cerebral infarct volume in MCAO rats, improves neurological function scores and pathological damage, which may be related to reducing the expression of IL-1b, IL-6, TNF-α and MCP-1 by downregulating TLR4, MyD88 and TRAF6 and reducing the expression and nuclear translocation of NF-κB. In addition, PTH can also reduce the levels of p-ERK1/2 to exert neuroprotective effect after ischemia (Zhang et al., 2021). Fasudil inhibits the activation of TLR4/NF-κB signal pathway and inflammatory response, exerting a neuroprotective effect (Zhang H. et al., 2018). In addition, Enriched environment (EE) is a method of exogenous intervention on the nervous system through behavioral intervention, which promotes neurogenesis and functional recovery through NF-kB-mediated secretion of IL-17A by Ast in stroke patients (Zhang Y. et al., 2018).