Bidirectional control of arteriole diameter by astrocytes

Astrocytes are the most numerous cells in the CNS. It is a defining feature of brain anatomy that every astrocyte has at least one contact with the vasculature, termed an endfoot. Collectively, all endfeet completely circumscribe all vessels in the brain. This unique anatomical feature has profound functional significance, as astrocyte endfeet have been discovered to release diffusible messengers that communicate directly with underlying smooth muscle cells to change arterial diameter and thereby regulate cerebral blood flow. A growing body of data now demonstrates that astrocytes serve as a bridge, relaying information on the level of neural activity to blood vessels in order to co-ordinate oxygen and glucose delivery with the energy demands of the tissue. In particular, astrocytes respond to elevations in extracellular glutamate as a consequence of synaptic transmission through the activation of group 1 metabotropic glutamate receptors. These Gq-coupled receptors elevate intracellular calcium via IP(3) signalling, which activates phospholipase A2 and generates arachidonic acid. Arachidonic acid acts as a signalling molecule or is converted to several lipid derivates, including prostaglandin E(2) and epoxyeicosatrienoic acids. Each of these lipids acts on vascular smooth muscle cells via different mechanisms to affect vessel diameter. Arachidonic acid initiates the production of 20-hydroxyeicosatetraenoic acid to cause vasoconstriction, whereas prostaglandin E(2) and epoxyeicosatrienoic acids cause vasodilatation. Factors that determine whether constrictor or dilatory pathways predominate involve nitric oxide and brain metabolic elements, such as oxygen, lactate and adenosine. Thus, astrocytes are thought to be capable of bidirectional control of arterial diameter, and the type of influence depends on the state of brain activity.     

Linear coupling between functional magnetic resonance imaging and evoked potential amplitude in human somatosensory cortex

The interpretation of task-induced functional imaging of the brain is critically dependent on understanding the relationship between observed blood flow responses and the underlying neuronal changes. However, the exact nature of this neurovascular coupling relationship remains unknown. In particular, it is unclear whether blood oxygen level-dependent functional magnetic resonance imaging (BOLD fMRI) responses principally reflect neuronal synaptic activity. In order to address this issue directly in humans, we measured the increase in somatosensory evoked potential amplitude and fMRI BOLD changes to increases in intensity of median nerve electrical stimulation in five healthy non-anaesthetized subjects. We found that mean N20-P22 amplitudes increased significantly with stimulus intensity in all subjects, as did fMRI BOLD percentage signal intensity change. Moreover, the intensity of the BOLD signal was found to correlate linearly with evoked potential amplitude in four of the five subjects studied. This suggests that the BOLD response correlates with synchronized synaptic activity, which is the major energy consuming process of the cortex.     

Neurovascular coupling in the mammalian brain

Normal brain function requires proper supply of oxygen and glucose in a timely and local manner. This is achieved through an orchestrated intercellular communication between neurones, astrocytes and microvessels that results in a rapid and restricted increase in cerebral blood flow, a process known as neurovascular coupling. Astrocytic end-feet make close contacts with neuronal synapses and blood vessels and, given their ability to release vasoactive signals following neuronal activation, have been recognized as key intermediaries in the neurovascular response. Both dilating and constricting signals appear to be released from astrocytes upon increases in intracellular Ca(2+) concentration, and both dilatation and constriction of brain vessels have been observed in previous studies. In this article, we discuss the various astrocyte-derived vasodilating and vasoconstricting signals, their interactions and effects on astrocytes and vascular smooth muscle cells, and suggest the importance of the intrinsic properties of the latter cell type on the overall neurovascular response. We present a working model in which the rise in astrocytic Ca(2+) following neuronal activation leads not only to the rapid activation of calcium-activated K(+) channels in astrocytic end-feet, but also to their modulation by metabolites of the arachidonic acid pathway, which in general have been proposed to act on vascular smooth muscle cells rather than on astrocytes. We propose that this latter mechanism may in turn modulate K(+) signalling from astrocytes to smooth muscle cells, influencing the overall effects of the vasodilating and vasoconstricting signals released during neuronal activation.     

Vascular amyloid alters astrocytic water and potassium channels in mouse models and humans with Alzheimer's disease

The neurovascular unit (NVU) comprises cerebral blood vessels and surrounding astrocytes, neurons, perivascular microglia and pericytes. Astrocytes associated with the NVU are responsible for maintaining cerebral blood flow and ionic and osmotic balances in the brain. A significant proportion of individuals with Alzheimer's disease (AD) have vascular amyloid deposits (cerebral amyloid angiopathy, CAA) that contribute to the heterogeneous nature of the disease. To determine whether NVU astrocytes are affected by the accumulation of amyloid at cerebral blood vessels we examined astrocytic markers in four transgenic mouse models of amyloid deposition. These mouse models represent mild CAA, moderate CAA with disease progression to tau pathology and neuron loss, severe CAA and severe CAA with disease progression to tau pathology and neuron loss. We found that CAA and disease progression both resulted in distinct NVU astrocytic changes. CAA causes a loss of apparent glial fibrillary acidic protein (GFAP)–positive astrocytic end-feet and loss of water channels (aquaporin 4) localized to astrocytic end feet. The potassium channels Kir4.1, an inward rectifying potassium channel, and BK, a calcium-sensitive large-conductance potassium channel, were also lost. The anchoring protein, dystrophin 1, is common to these channels and was reduced in association with CAA. Disease progression was associated with a phenotypic switch in astrocytes indicated by a loss of GFAP-positive cells and a gain of S100β-positive cells. Aquaporin 4, Kir4.1 and dystrophin 1 were also reduced in autopsied brain tissue from individuals with AD that also display moderate and severe CAA. Together, these data suggest that damage to the neurovascular unit may be a factor in the pathogenesis of Alzheimer's disease.     

Evidence that angiogenesis lags behind neuron and astrocyte growth in experience-dependent plasticity

Bill Greenough's work on the cell biology of information storage suggests that we cannot understand the mechanism of long-term memory without understanding the series of cellular transactions that drive coordinated structural changes in neurons, glia, and blood vessels. Here, we show that after 4 days of differential housing, neuropil of EC cortex has expanded significantly, but the vasculature has not, resulting in a dilution of the blood supply. Significant growth of neurons and astrocytes has been reported within this time period, suggesting expression of synaptic plasticity might involve temporally coordinated genomic responses by both neurons and glia. Given that astrocytes appear to couple neuronal and vascular growth during development, we hypothesize that they may also mediate the onset of angiogenesis in response to neural demand in the EC brain. Further, these results may imply that a neuron's capacity for plasticity could be constrained by the rate of vascular expansion.     

The neurovascular unit in brain function and disease

Neurovascular coupling, or functional hyperaemia, refers to complex mechanisms of communication between neurons, astrocytes and cerebral vessels which form the neurovascular unit that spatially and temporally adjusts blood supply to the needs in energy and oxygen of activated neurons. Neurovascular coupling is so precise that it underlies neuroimaging techniques to map changes in neuronal activity. Therefore, understanding its basis is indispensable for the proper interpretation of imaging signals from functional magnetic resonance imaging and positron emission tomography, routinely used in humans. Although neurovascular coupling mechanisms are not yet fully understood, considerable progress has been made over the last decade. In this review, we present recent knowledge from in vivo studies on the cortical cellular network involved in neurovascular coupling responses and the mediators implicated in these haemodynamic changes. Recent findings have emphasized the intricate interplay between both excitatory and inhibitory neurons in neurovascular coupling, together with an intermediary role of astrocytes, which are ideally positioned between neurons and microvessels. Finally, we describe latest findings on the alterations of neurovascular function encountered in neurodegenerative conditions such as Alzheimer's disease.     

Molecular mechanisms controlling nutritive blood flow: role of cytochrome P450 enzymes

This short review summarizes the potential role of cytochrome P450 (P450) in regulating blood flow in the brain tissue and in the skeletal muscle. We provide data showing that pressure-induced myogenic activity in the brain is largely responsible for autoregulation of CBF. This myogenic response to pressure is maintained, in part, by 20-HETE formation in arterial muscle cells through a P450 omega-hydroxylase coded for by a P450 4A cDNA. Autoregulation of CBF is a hallmark of the cerebral circulation and provides adequate nutritive blood flow despite large fluctuations in arterial pressure. Given the importance of oxidative metabolism in the brain, support of neuronal activity is mediated by functional hyperaemia to active neurones providing adequate delivery of oxidative substrate. We provide data demonstrating that this functional hyperaemia in the brain is regulated by astrocytes which sense neural activity and release dilator metabolites which shunt blood flow to active neurones. One of the metabolites released by astrocytes in this regard are epoxygenated products of arachidonic acid (AA) formed by P450 enzymes. These AA metabolites of P450 enzymes are epoxyeicosatrienoic acid (EETs). One of these P450 enzymes is coded by a 2C11 cDNA present in astrocytes. Furthermore, astrocytes are capable of inducing capillary angiogenesis which appears to be mediated, in part, by P450-derived EETs.     

Signalling within the neurovascular unit in the mammalian retina

Neuronal activity in the central nervous system evokes localized changes in blood flow, a response termed neurovascular coupling or functional hyperaemia. Modern functional imaging methods, such as functional magnetic resonance imaging (fMRI), measure signals related to functional hyperaemia in order to determine localization of brain function and to diagnose disease. The cellular mechanisms that underlie functional hyperaemia, however, are not well understood. Glial cells have been hypothesized to be intermediaries between neurons and blood vessels in the control of neurovascular coupling, owing to their ability to release vasoactive factors in response to neuronal activity. Using an in vitro preparation of the isolated, intact rodent retina, we have investigated two likely mechanisms of glial control of the vasculature: glial K(+) siphoning and glial induction of vasoactive arachidonic acid metabolites. Potassium siphoning is a process by which a K(+) current flowing through glial cells transfers K(+) released from active neurons to blood vessels. Since slight increases in extracellular K(+) can cause vasodilatation, this mechanism was hypothesized to contribute to neurovascular coupling. Our data, however, suggest that glial K(+) siphoning does not contribute significantly to neurovascular coupling in the retina. Instead, we suggest that glial cells mediate neurovascular coupling by inducing the production of two types of arachidonic acid metabolites, epoxyeicosatrienoic acids (EETs) and 20-hydroxyeicosatetraenoic acid (20-HETE), which dilate and constrict vessels, respectively. We show that both light flashes and direct glial stimulation produce vasodilatation or vasoconstriction mediated by EETs and 20-HETE, respectively. Further, we show that the type of vasomotor response observed (dilatation or constriction) depends on retinal levels of nitric oxide. Our data also demonstrate that glial cells are necessary intermediaries for signalling from neurons to blood vessels, since functional hyperaemia does not occur when neuron-to-glia communication is interrupted. These results indicate that glial cells play an important role in mediating functional hyperaemia and suggest that the regulation of blood flow may involve both vasodilating and vasoconstricting components.     

The Pericyte: A Forgotten Cell Type with Important Implications for Alzheimer's Disease?

Pericytes are cells in the blood–brain barrier (BBB) that degenerate in Alzheimer's disease (AD), a neurodegenerative disorder characterized by early neurovascular dysfunction, elevation of amyloid β‐peptide (Aβ), tau pathology and neuronal loss, leading to progressive cognitive decline and dementia. Pericytes are uniquely positioned within the neurovascular unit between endothelial cells of brain capillaries, astrocytes and neurons. Recent studies have shown that pericytes regulate key neurovascular functions including BBB formation and maintenance, vascular stability and angioarchitecture, regulation of capillary blood flow, and clearance of toxic cellular by‐products necessary for normal functioning of the central nervous system (CNS). Here, we review the concept of the neurovascular unit and neurovascular functions of CNS pericytes. Next, we discuss vascular contributions to AD and review new roles of pericytes in the pathogenesis of AD such as vascular‐mediated Aβ‐independent neurodegeneration, regulation of Aβ clearance and contributions to tau pathology, neuronal loss and cognitive decline. We conclude that future studies should focus on molecular mechanisms and pathways underlying aberrant signal transduction between pericytes and its neighboring cells within the neurovascular unit, that is, endothelial cells, astrocytes and neurons, which could represent potential therapeutic targets to control pericyte degeneration in AD and the resulting secondary vascular and neuronal degeneration.     

Principal neuron spiking: neither necessary nor sufficient for cerebral blood flow in rat cerebellum

Neuronal activity, cerebral blood flow, and metabolic responses are all strongly coupled, although the mechanisms behind the coupling remain unclear. One of the key questions is whether or not increases in spiking activity in the stimulated neurons are sufficient to drive the activity-dependent rises in cerebral blood flow (CBF) that form the basis of the signals used in functional neuroimaging such as the blood oxygen level-dependent (BOLD) signal. To this end the present study examined the effect of enhanced spike activity per se on CBF in rat cerebellar cortex under conditions of disinhibition, achieved by blocking GABA_A receptors using either bicuculline or picrotoxin. Purkinje cell spiking activity and local field potentials were recorded by glass microelectrodes, and laser Doppler flowmetry was used to monitor CBF. Disinhibition increased Purkinje cell spiking rate to 200–300% of control without incurring any increase in basal CBF. This demonstrates that increased spike activity per se is not sufficient to affect basal CBF. The neurovascular coupling between excitatory synaptic activity and CBF responses evoked by inferior olive (climbing fibre) stimulation was preserved during disinhibition. Thus, the unchanged basal CBF in the presence of the dramatic rise in Purkinje cell spiking rate was not explained by impaired synaptic activity–CBF coupling. On the basis of our previous and the present studies, we conclude that increased spiking activity of principal neurons is neither sufficient nor necessary to elicit CBF responses and in turn BOLD signals, and that activation-dependent vascular signals reflect excitatory synaptic activity.     

Astrocyte-mediated control of cerebral blood flow

Local increase in blood flow during neural activity forms the basis for functional brain imaging, but its mechanism remains poorly defined. Here we show that cortical astrocytes in vivo possess a powerful mechanism for rapid vasodilation. We imaged the activity of astrocytes labeled with the calcium (Ca(2+))-sensitive indicator rhod-2 in somatosensory cortex of adult mice. Photolysis of caged Ca(2+) in astrocytic endfeet ensheathing the vessel wall was associated with an 18% increase in arterial cross-section area that corresponded to a 37% increase in blood flow. Vasodilation occurred with a latency of only 1-2 s, and both indomethacin and the cyclooxygenase-1 inhibitor SC-560 blocked the photolysis-induced hyperemia. These observations implicate astrocytes in the control of local microcirculation and suggest that one of their physiological roles is to mediate vasodilation in response to increased neural activity.     

The neurobiology of glia in the context of water and ion homeostasis

Astrocytes are highly complex cells that respond to a variety of external stimulations. One of the chief functions of astrocytes is to optimize the interstitial space for synaptic transmission by tight control of water and ionic homeostasis. Several lines of work have, over the past decade, expanded the role of astrocytes and it is now clear that astrocytes are active participants in the tri-partite synapse and modulate synaptic activity in hippocampus, cortex, and hypothalamus. Thus, the emerging concept of astrocytes includes both supportive functions as well as active modulation of neuronal output. Glutamate plays a central role in astrocytic-neuronal interactions. This excitatory amino acid is cleared from the neuronal synapses by astrocytes via glutamate transporters, and is converted into glutamine, which is released and in turn taken up by neurons. Furthermore, metabotropic glutamate receptor activation on astrocytes triggers via increases in cytosolic Ca(2+) a variety of responses. For example, calcium-dependent glutamate release from the astrocytes modulates the activity of both excitatory and inhibitory synapses. In vivo studies have identified the astrocytic end-foot processes enveloping the vessel walls as the center for astrocytic Ca(2+) signaling and it is possible that Ca(2+) signaling events in the cellular component of the blood-brain barrier are instrumental in modulation of local blood flow as well as substrate transport. The hormonal regulation of water and ionic homeostasis is achieved by the opposing effects of vasopressin and atrial natriuretic peptide on astroglial water and chloride uptake. In conjuncture, the brain appears to have a distinct astrocytic perivascular system, involving several potassium channels as well as aquaporin 4, a membrane water channel, which has been localized to astrocytic endfeet and mediate water fluxes within the brain. The multitask functions of astrocytes are essential for higher brain function. One of the major challenges for future studies is to link receptor-mediated signaling events in astrocytes to their roles in metabolism, ion, and water homeostasis.     

Cortical spreading depression produces long-term disruption of activity-related changes in cerebral blood volume and neurovascular coupling

Cortical spreading depression (CSD) is a pronounced depolarization of neurons and glia that spreads slowly across the cortex followed by a period of depressed electrophysiological activity. The vascular changes associated with CSD are a large transient increase in blood flow followed by a prolonged decrease lasting greater than 1 h. Currently, the profile of functional vascular activity during this hypovolemic period has not been well characterized. Perfusion-based imaging techniques such as functional magnetic resonance imaging (fMRI) assume a tight coupling between changes in neuronal and vascular activity. Under normal conditions, these variables are well correlated. Characterizing the effect of CSD on this relationship is an important step to understand the impact acute pathophysiological events may have on neurovascular coupling. We examine the effect of CSD on functional changes in cerebral blood volume (CBV) evoked by cortical electrophysiological activity for 1 h following CSD induction. CBV signal amplitude, duration, and time to peak show little recovery at 60 min post-induction. Analysis of spontaneous vasomotor activity suggests a decrease in vascular reactivity may play a significant role in the disruption of normal functional CBV responses. Electrophysiological activity is also attenuated but to a lesser degree. CBV and evoked potentials are not well correlated following CSD, suggesting a breakdown of the neurovascular coupling relationship.     

Msx1 expression in the adult mouse brain: characterization of populations of beta-galactosidase-positive cells in the hippocampus and fimbria

We have analyzed Msx1 expression in the mature mouse brain using in situ hybridization and beta-galactosidase activity in Msx1(nLacZ) mice. The study revealed that Msx1 is strongly expressed in the circumventricular organs, such as the subcommissural organ and choroid plexus, and in some epithelia, such as that of the dorsal, but not the ventral part of the third ventricle. Immunohistochemical analysis revealed that the Msx1-expressing cells of the hippocampus and fimbria are astrocytes, oligodendrocytes or immature oligodendrocytes. In contrast, no co-expression was detected in these structures using several neuronal markers. These results were confirmed, using transmission electron microscopy, by the presence of 5-bromo-3-indolyl-beta-D-galactopyranosideprecipitates in astrocytes and oligodendrocytes in both sites. Moreover, using an anti-glial fibrillary acidic protein antibody (GFAP), our study reveals two populations of astrocytes in the adult hippocampus and other areas, such as the fimbria, namely Msx1+/GFAP+ and Msx1-/GFAP+. Beta-galactosidase activity was also observed in endothelial cells of hippocampal fissure blood vessels. We also observed co-localization of polysialic acid neural cell adhesion molecule, a marker of the polysialylated form of the neural cell adhesion molecule, in Msx1-expressing cells in the fimbria. These cells may be precursors of glial cells and originate from the epithelium of the fimbria. The present study indicates, in the mature mouse brain, that Msx1 may be linked to secretory activity in circumventricular organs, and to glial proliferation and differentiation in the hippocampus and fimbria, and presumably also in other cerebral areas. We suggest that Msx1 could be associated with brain homeostasis and blood-brain barrier function.     

Astrocytes in a dish: Using pluripotent stem cells to model neurodegenerative and neurodevelopmental disorders

Neuro science and Neuro biology have historically been neuron biased, yet up to 40% of the cells in the brain are astrocytes. These cells are heterogeneous and regionally diverse but universally essential for brain homeostasis. Astrocytes regulate synaptic transmission as part of the tripartite synapse, provide metabolic and neurotrophic support, recycle neurotransmitters, modulate blood flow and brain blood barrier permeability and are implicated in the mechanisms of neurodegeneration. Using pluripotent stem cells (PSC), it is now possible to study regionalised human astrocytes in a dish and to model their contribution to neurodevelopmental and neurodegenerative disorders. The evidence challenging the traditional neuron‐centric view of degeneration within the CNS is reviewed here, with focus on recent findings and disease phenotypes from human PSC‐derived astrocytes. In addition we compare current protocols for the generation of regionalised astrocytes and how these can be further refined by our growing knowledge of neurodevelopment. We conclude by proposing a functional and phenotypical characterisation of PSC‐derived astrocytic cultures that is critical for reproducible and robust disease modelling.     

内皮细胞对神经干细胞增殖与分化的影响

已经确定神经干细胞在脑中不是随意分布,而是集中在血管周围的。尽管神经干细胞存在于血管周围,但是对于其与血管组成细胞之间的关系还不是很清楚。据报道,内皮细胞释放的可溶性因子可以刺激神经干细胞自我增殖,抑制其分化,并且提高神经元的比例。将内皮细胞与神经干细胞共同培养可以激活Notch途径来促进神经干细胞自我增殖。另外,血管内皮生长因子对神经细胞的生长也起着非常重要的作用,它促进了中枢神经系统星形胶质细胞的生长与分化。因此,内皮细胞不仅是传统意义上血管的组成部分,还是神经干细胞所在区域的重要成分,并且可以通过大脑产生的神经营养性分泌物来提高神经元的发生。     

Synaptic modulation by astrocytes via Ca2+-dependent glutamate release

In the past 15 years the classical view that astrocytes play a relatively passive role in brain function has been overturned and it has become increasingly clear that signaling between neurons and astrocytes may play a crucial role in the information processing that the brain carries out. This new view stems from two seminal observations made in the early 1990s: 1. astrocytes respond to neurotransmitters released during synaptic activity with elevation of their intracellular Ca2+ concentration ([Ca2+]i); 2. astrocytes release chemical transmitters, including glutamate, in response to [Ca2+]i elevations. The simultaneous recognition that astrocytes sense neuronal activity and release neuroactive agents has been instrumental for understanding previously unknown roles of these cells in the control of synapse formation, function and plasticity. These findings open a conceptual revolution, leading to rethink how brain communication works, as they imply that information travels (and is processed) not just in the neuronal circuitry but in an expanded neuron-glia network. In this review we critically discuss the available information concerning: 1. the characteristics of the astrocytic Ca2+ responses to synaptic activity; 2. the basis of Ca2+-dependent glutamate exocytosis from astrocytes; 3. the modes of action of astrocytic glutamate on synaptic function.     

High-resolution neurometabolic coupling revealed by focal activation of visual neurons

Functional magnetic resonance imaging is an important tool for measuring brain function noninvasively, but the vascular and metabolic changes on which its measurements are based are not fully understood. Here, we examined the relationship between these changes and neural activity on a fine spatial scale through simultaneous measurements of tissue oxygen and extracellular neural activity in the cat lateral geniculate nucleus. Our findings indicate that activity-dependent increases in cerebral blood flow and oxidative metabolism occur on different spatial scales, and that the ratio between the two depends on the size of the activated neural population.     

Synaptic transmission and plasticity are modulated by nonmuscle myosin II at the neuromuscular junction of Drosophila

The synaptic vesicle population in a nerve terminal is traditionally divided into subpopulations according to physiological criteria; the readily releasable pool (RRP), the recycling pool, and the reserve pool. It is recognized that the RRP subserves synaptic transmission evoked by low-frequency neural activity and that the recycling and reserve populations are called on to supply vesicles as neural activity increases. Here we investigated the contribution of nonmuscle myosin II (NMMII) to synaptic transmission with emphasis on the role a motor protein could play in the supply of vesicles. We used Drosophila genetics to manipulate NMMII and assessed synaptic transmission at the larval neuromuscular junction. We observed a positive correlation between synaptic strength at low-frequency stimulation and NMMII expression: reducing NMMII reduced the evoked response, while increasing NMMII increased the evoked response. Further, we found that NMMII contributed to the spontaneous release of vesicles differentially from evoked release, suggesting differential contribution to these two release mechanisms. By measuring synaptic responses under conditions of differing external calcium concentration in saline, we found that NMMII is important for normal synaptic transmission under high-frequency stimulation. This research identifies diverse functions for NMMII in synaptic transmission and suggests that this motor protein is an active contributor to the physiology of synaptic vesicle recruitment.     

ATP release and hydrolysis contribute to rat pial arteriolar dilatation elicited by neuronal activation

Owing to their intimate anatomical relationship with cerebral arterioles, astrocytes have been postulated as signal transducers, transferring information from activated neurones to the cerebral microcirculation. These forwarded signals may involve the release of vasoactive factors from the end-feet of astrocytes. This mechanism is termed 'neurovascular coupling' and its anatomical components (i.e. neurone, astrocyte and vascular cells) are termed the 'neurovascular unit'. The process of neurovascular coupling often involves upstream dilatation. This is necessary during periods of increased metabolic demand, in order to permit more blood to reach dilated downstream vessels, thereby improving nutrient supply to the activated neurones. Without it, that downstream dilatation might be ineffective, placing neurones at risk, especially during episodes of intense neuronal activity, such as seizure. In the brain, pial arterioles represent important 'upstream' vascular segments. The pial arterioles overlie a thick layer of astrocytic processes, termed the glia limitans. This essentially isolates pial arterioles, anatomically, from the neurones below. Vasodilating signals that originate in the neurones therefore reach the pial arterioles via indirect pathways, primarily involving astrocytes and the glia limitans. Here we discuss a process whereby purinergic mechanisms play a key and neuronal activity-dependent role in astrocyte to astrocyte communication, as well as in glia limitans to pial arteriolar signals leading to vasodilatation.