Well shit your doctor should have been figuring out how to prevent pericytes from strangling capillaries for years. Did nothing, why are you seeing them?
Capillaries that don't open due to pericytes September 2011
So only 12 years for your incompetent doctor do do nothing on pericytes!
Pericytes and the Control of Blood Flow in Brain and Heart
Annual Review of Physiology
Vol. 85:137-164 (Volume publication date February 2023)
https://doi.org/10.1146/annurev-physiol-031522-034807
Abstract
Pericytes, attached to the surface of capillaries, play an important role in regulating local blood flow. Using optogenetic tools and genetically encoded reporters in conjunction with confocal and multiphoton imaging techniques, the 3D structure, anatomical organization, and physiology of pericytes have recently been the subject of detailed examination. This work has revealed novel functions of pericytes and morphological features such as tunneling nanotubes in brain and tunneling microtubes in heart. Here, we discuss the state of our current understanding of the roles of pericytes in blood flow control in brain and heart, where functions may differ due to the distinct spatiotemporal metabolic requirements of these tissues. We also outline the novel concept of electro-metabolic signaling, a universal mechanistic framework that links tissue metabolic state with blood flow regulation by pericytes and vascular smooth muscle cells, with capillary KATP and Kir2.1 channels as primary sensors. Finally, we present major unresolved questions and outline how they can be addressed.
Keywords
pericytes, heart, brain, metabolism, neurovascular coupling, electro-metabolic signaling, ion channels, calcium, potassium
INTRODUCTION
Pericytes were first observed and documented by Eberth (1) and Rouget (2) approximately 150 years ago, and the term pericyte (surrounding cell, from the Greek prefix peri- denoting something that is around or nearby) was coined by Zimmermann a century ago (3). Even without modern imaging methods, the illustrations in these early works were able to capture the beauty and diversity of pericyte morphology with stunning detail and accuracy (Figure 1a). This diversity has both intrigued and confounded physiologists attempting to understand pericyte function since their discovery and continues to be a source of considerable controversy. However, advances in genetic engineering, imaging, and molecular profiling are now enabling studies that probe the functions of the cells that compose the pericyte continuum with unprecedented depth and precision, and the recent exponential increase in interest in pericyte biology promises to rapidly advance our future understanding.
The provocative positioning of pericytes on the outer wall of capillaries—their processes and cell bodies making intimate contact with adjacent capillary endothelial cells (cECs)—prompted early speculation that they play roles in controlling blood flow (4). In the intervening years, mounting evidence has confirmed this, although we still lack a detailed mechanistic understanding of these processes. Studying the functions of pericytes in organ systems with distinct metabolic requirements and different blood flow profiles may not only underscore commonalities of pericyte structure and function but also provide opportunities to uncover specializations that have evolved to equip these cells with the ability to regulate blood flow according to specific local energy demands. Accordingly, here we focus our attention on the emerging roles of pericytes in blood flow control in both heart and brain in health and disease.
As the primary pump driving blood around the body, the heart must work consistently throughout an animal's lifetime to ensure uninterrupted delivery of oxygen (O2) and nutrients. Cardiac cells beat continuously, with their pace—and therefore energy requirements—modulated by external factors such as sympathetic:parasympathetic balance, hormone levels, and overwhelmingly the degree of whole-animal activity (5–8). Intrinsic local control of cardiac blood flow operates under the control of an important electro-metabolic signaling (EMS) mechanism, first identified in our recent work and in which pericytes play a vital role that is in the process of being defined (9, 10). The brain, in contrast to heart, exhibits variations in activity and energy demand across broader spatiotemporal scales, dictated by the computational demands that are placed upon neuronal networks spanning local and interregional territories at any given time. Blood flow in this context is continually adjusted to meet these highly fluctuating energy requirements through the process of neurovascular coupling (11, 12), where the cells of the vasculature modulate blood flow via the engagement of local signaling cascades in response to increased neural activity. A primary form of local blood flow control in brain is exerted by parenchymal potassium concentration ([K+]), which fluctuates according to neuronal activity (13). K+ controls the activity of the strong inward rectifier K+ channel, Kir2.1, which plays an important functional role in cECs (14, 15). In both heart (10) and brain (14, 15), this feature of cECs lays the foundation for rapid electrical signaling throughout the vasculature to regulate blood flow. A critically important emerging feature of both heart and brain pericyte physiology is that these cells can communicate electrically through gap junctions with underlying cECs to modulate this ongoing electrical activity (9, 10, 16–19).
Recent reviews (20, 21) have focused broadly and extensively on the multitude of molecular mechanisms through which pericytes may control blood flow. Here, our focus is on what can be learned from pericytes operating in two different organ systems with high energy requirements, and in particular, their potential roles in energy sensing and electrical signaling to control blood flow. We begin by reviewing the challenges associated with blood flow control in brain and heart and then assess the morphological and functional features of pericytes in these organs. We then focus on understanding the roles for pericytes in supporting neuronal and myocyte function through precision modulation of blood flow in these distinct contexts.
CONTROL OF BLOOD FLOW IN BRAIN AND HEART
The angioarchitecture of the cerebral cortex consists of pial arteries on the brain's surface, which branch at right angles, yielding functionally distinct penetrating arterioles that dive into the tissue (Figure 1). From here, an incredibly dense and tortuous capillary bed emerges with pericytes and their processes adorning most of the outer wall of this network. Given that the capillary bed composes ∼90% of the brain vasculature by volume (22) and that these vessels are tightly interwoven with neuronal processes and cell bodies, the pericytes found here are ideally positioned to receive and process information on local neuronal activity and tune blood flow in response. Depending on their form and location within the capillary bed (see below), pericytes make differing contributions to this processing and help to solve several key challenges for blood delivery throughout the brain.
At the global level, the fixed volume of the skull imposes limitations on vascular dynamics and as such substantial changes in the volume of blood or cerebrospinal fluid can affect intracranial pressure and damage the brain (23, 24). Thus, changing global perfusion to the brain by dilating extensive portions of major feed arteries and arterioles (e.g., as might happen in working muscle) may adversely affect intracranial pressure and endanger neuronal function and therefore needs to be avoided. Consequently, the brain has evolved intricate mechanisms to focally modulate vessel diameter and blood flow, avoiding potentially damaging large changes in perfusion. It is now emerging that pericytes play a major role in this process.
At the local level, neuronal activity varies dramatically within individual cells and across populations. For example, pyramidal neurons discharge at about 1 Hz at rest and may increase their firing rate by more than an order of magnitude during activity (25–28), whereas fast-spiking interneurons from humans can fire in excess of 300 Hz (29). Increases in firing rate impose a huge increase in energy consumption (30), and the energy required to underpin this activity is primarily dedicated to ionic pumping mechanisms such as the sodium/potassium (Na+/K+) ATPase and the plasma membrane calcium (Ca2+) ATPase, which reverse the ionic fluxes that occur during action potentials and synaptic activity (30, 31). As the energy needed to fuel these processes is ultimately derived from the blood, this necessitates tight functional linkage between neuronal activity and increases in blood flow through the process of functional hyperemia, underpinned by the mechanisms of neurovascular coupling. This blood flow increase may also serve other important homeostatic functions, such as waste removal and temperature regulation. Accordingly, the brain has evolved a series of unique, layered, and redundant blood flow control mechanisms that span varied spatiotemporal scales and operate in concert to guarantee energy supply through orchestrated molecular activity that plays out across the multiple cell types of the neurovascular unit [endothelial cells (ECs) and smooth muscle cells (SMCs), pericytes, neurons, and astrocytes] (32). Pericytes sit at the center of this activity, nestled between astrocytic endfeet and the EC wall, and are thus ideally positioned to facilitate communication between the parenchyma and the vasculature to regulate local blood flow (33).
A distinct set of blood flow control problems accompany cardiac activity, and coronary blood flow control mechanisms (i.e., regulation of blood flow that the heart supplies to itself through the coronary arteries) have evolved to guarantee a continuous and extremely efficient supply of energy to support moment-to-moment heart work for an entire lifetime. Here too, it is now emerging that pericytes are intimately involved in these processes. Coronary blood flow varies with time depending on aortic pressure, myocardial extravascular pressure, and resistance to flow, which, in turn, are critically dependent on myocardial metabolism and neural and hormonal controls (for reviews, see 5, 7, 34, 35). These influences converge to regulate the contractile state of the small arterioles of the heart and their SMCs. The functional unit of blood flow regulation in heart is shown in Figure 3e. This functional unit embodies three sets of components that form the EMS system in heart. First, the ventricular myocytes are the primary metabolic sensors; second, the capillary pericytes, contractile pericytes, and end-arteriole smooth muscle cells are the primary blood flow regulators; and third, the local cECs are the primary electrical transmission elements in the local signal distribution network (10). We focus here on small vessel regulation in heart, as these vessels are more likely to receive direct input from local downstream pericytes and signals from adjacent ventricular myocytes, but we note that larger vessels also contribute to this process (35).
Coronary blood flow must be nearly perfectly matched to the metabolic demands of the heart at all times, which is evidenced by a near-maximal extraction of O2 from blood in the heart under all flow conditions. Blood appears to be directed to where there is metabolic need and is diverted away from regions that are replete with nutrients. The heart will adjust flow partially to compensate for local deficits where appropriate, a system that is somewhat analogous to control of blood flow in the brain by neurovascular coupling. However, the amount of O2 consumed by the heart and extracted from coronary blood is the highest of any organ per gram of tissue (36, 37). Put another way, the arterio-venous pO2 (partial pressure of oxygen) difference in heart is nearly always maximal, and the ratio of arterial pO2 to venous pO2 is maintained at around 1.6 to 1.7 unless the arterial pO2 falls to under 60% of its normal level (36, 37). It appears that a combination of the voltage sensitivity of SMC contractions, the electrical and contractile functions of pericytes, and their local signaling functions must work in tandem to continually and dynamically adjust blood flow through this system over a wide range of conditions. Nevertheless, precise, quantitative experiments are needed to accurately determine how pericytes in both of these contexts work, how they collate and distribute information, and how they act on it to control blood flow.
THE PERICYTE CONTINUUM
Ongoing debates surrounding the precise identity and classification of pericytes throughout the capillary bed require that we define both the varieties of these cells at the different levels of the vascular tree and in different organ systems and the criteria used to delineate the border between arterioles and capillaries. In the brain, extensive and ongoing work has been performed to carefully identify the morphological, functional and transcriptomic properties of pericytes throughout the vascular bed (Figure 2f). Arteries and arterioles can be distinguished from capillaries by two major features. (a) Arteries and arterioles secrete elastin (38, 39), a major component of the internal elastic lamina that can be stained to highlight these vessels. This elastin lining abruptly ends at the transition to the capillary bed (40) (Figure 1b). (b) Arteries and arterioles are suitably wide to allow the passage of red blood cells (RBCs) side by side, whereas capillaries are narrow enough to allow only for single-file RBC transit (Figure 1c). Accordingly, we define the initial capillary of a vascular network under consideration as the first to display a lack of elastin staining and to permit RBC passage in single file. Brain capillaries are referred to by branch order, so that the first capillary emerging from the arteriole is labeled as the first-order capillary. This numbering increases by one each time the capillaries branch as they extend deeper into the parenchyma, regardless of vessel diameter or orientation (Figure 1d). In the heart, the size and biochemical distinctions noted above hold true, but the ordered, sequential branching of capillaries rapidly breaks down as one examines the deeper capillary bed. Instead, capillaries here are much more elaborately and abundantly interconnected, with frequent anastomoses between vessels arrayed in near-parallel fashion around adjacent cardiac myocytes to form a looping, meshed network through which RBCs may take many potential paths (10, 41–43) (Figure 1e,f). In both systems, the capillary bed terminates when vessels unify at venules that pass deoxygenated blood and waste products out to larger veins (Figure 1d,e).
Arteries and arterioles are covered by SMCs that receive local electrical signals from their underlying arteriolar ECs and from downstream cECs. These SMCs enwrap the underlying cobblestone (44) arteriolar EC network concentrically, and the mechanisms that underlie their regulation have been extensively studied (Supplemental Box 1). SMCs are distinguished from bump-on-a-log, undulating pericytes by their concentric orientation and relatively flat, banded, fusiform morphology (45–48).
At the transition to the capillary bed, a specialized structure known as the precapillary sphincter has been identified, present at ∼70% of all such branch points (49). Control of this juncture and the initial branches that follow is key to regulating perfusion of the capillary bed, mediated through the dynamic regulation of the contractile state of α-smooth muscle actin (α-SMA)-positive mural cells (49–51). Indeed, in the brain the first few capillary branches are covered by such contractile cells that actively regulate the diameter of the underlying vessel, which have been variably referred to as contractile pericytes (48, 52), SMCs (45, 53), ensheathing pericytes (20, 54), and hybrid cells (55). Here, we use the term contractile pericyte as, in our view, this best reflects the mounting evidence that these cells are functionally distinct from upstream SMCs, and this name helps to distinguish these cells from their less contractile counterparts further downstream. However, we emphasize that this is not an exclusive term. Rather, its use helps to underscore the key role of these cells in rapid diameter control of the underlying capillaries. Other pericyte types are also capable of contracting, but they appear to operate on slower timescales and have more subtle effects on diameter (56). Indeed, a major feature that contractile pericytes have in common with upstream SMCs is their expression of α-SMA (42, 48, 54, 57), and it is noteworthy that these cells cannot be distinguished from SMCs in genetic screens (58–60), likely as a result of their common lineage (61). However, mounting morphological and functional differences can be used to distinguish contractile pericytes from their SMC counterparts. Morphologically, contractile pericytes are distinguished by their bulging, ovoid cell bodies that give way to large finger-like projections that enwrap the underlying capillary (Figures 1a and 2b,f), covering up to about 95% of the outer surface of the vessel and giving the pericyte tight control over vessel diameter (46, 54). At the protein level, the Ca2+-binding protein calponin is detectable in SMCs but is not evident in contractile pericytes, and the same is true for polymerized microtubules (48). In contrast, contractile pericytes exhibit high levels of the intermediate filament desmin, aminopeptidase N (also known as CD13), the membrane chondroitin sulfate proteoglycan NG2, and platelet-derived growth factor receptor-β (PDGFRβ), which starkly contrasts with lower levels of these proteins in upstream SMCs (42, 57). Control of cytosolic Ca2+ is central to the regulation of the tone of the Ca2+-sensitive contractile apparatus in cardiac, skeletal, and smooth muscle (Supplemental Box 1). SMCs rely heavily on Ca2+ release through ryanodine receptors (RyRs) to regulate tone (62–64), whereas functional RyRs have not been detected in contractile pericytes (48, 65). Instead, inositol-1,4,5-trisphosphate (IP3) receptor–mediated intracellular Ca2+ signaling appears to play a prominent role in the contraction of these cells (48). It is anticipated that more differences between SMCs and contractile pericytes will emerge through further experimental efforts.
In the cortex and retina, the initial 3–4 branches of the capillaries emerging from the arteriole are covered by these contractile pericytes (Figure 2f,h), which play an important role in the branch-by-branch control of local blood flow. Immediately downstream of contractile pericytes on ∼fourth-order capillaries in brain is a population of mesh pericytes that can be distinguished on the basis of their morphology, possessing a tangled network of processes, covering a lower fraction of the capillary surface, and expressing lower levels of α-SMA (54). Little is known of these cells, and the fact that a distinct transcriptomic signature to separate them from adjacent pericyte types has not been identified makes finding inroads for further detailed study challenging (66). Downstream of mesh pericytes, from approximately the fifth branch order and beyond, the capillaries are covered by thin-strand pericytes (Figure 2f,h). These cells express very low levels of α-SMA and extend long, thin filamentous processes away from the cell body that reach in some cases for hundreds of microns (54, 67, 68), taking on a diverse range of shapes and lengths.
Owing to the high density of capillaries in heart [∼2,000–3,000 capillaries/mm2 (69)] and the distinct organization of both the capillaries and pericytes in this system, we use an alternate but related terminology to divide the pericytes found in heart into two types: contractile pericytes and capillary pericytes (42, 70–72). Based on our own and existing morphological observations using electron and confocal microscopy, we observe that α-SMA–containing pericytes are primarily seen at arteriolar-capillary junctions (see Figure 2b,e) in heart, which we term contractile pericytes. This name distinguishes them from the more abundant and again less contractile pericytes found deeper in the complex, anastomosing cardiac capillary bed, called capillary pericytes (42, 70, 71, 73, 74) (Figure 2a). We describe these subtypes in greater detail below. A complicating factor in drawing such a limited distinction is that cardiac pericytes lack cell- and subtype-specific biomarkers (a similar problem plagues the pericytes of the brain) that can be used for their unequivocal identification and classification. This issue highlights the need for an improved strategy for pericyte/SMC distinction, which might be achieved with a dual labeling system. For example, expressing one fluorophore under the control of the promoter of the elastin gene, Eln, and another fluorophore with separable spectral properties under the PDGFRβ gene, Pdgfrb, to distinguish elastin+/ PDGFRβ+ SMCs from elastin−/ PDGFRβ+ pericytes might be useful in this regard. However, in keeping with current strategies used to study brain pericytes, NG2 (encoded by the Cspg4 gene) and PDGFRβ (Pdgfrb) alone are two markers that can be used to at least initially identify cardiac pericytes and reveal their physical structures and locations (10, 42, 43, 60, 75–77).
Cardiac contractile pericytes wrap around the components of the arteriolar-capillary junction (78) (Figure 2b), a positioning that enables them to exert powerful influence over local blood flow. Both contractile and capillary pericytes in heart appear to be capable of contracting to differing degrees (42), which aligns with recent findings for brain capillary pericytes (48, 52, 56). In the brain, upstream contractile pericytes contract more robustly and more quickly than their downstream thin-strand counterparts (48, 56). However, pericyte contraction dynamics have not been as thoroughly characterized in heart, regardless of the findings on fixed and nonperfused tissues indicating that pericytes might be involved in capillary constriction in ischemia-reperfusion (42); this area awaits further detailed study.
Pericytes on deeper heart capillaries are broadly similar to the thin-strand pericytes of the brain, but the lack of a reliably sequential vascular branching pattern that leads to an easily recognizable capillary hierarchy makes their identification based on branch order much more difficult (50). Indeed, the ovoid cell body of heart pericytes protrudes from the capillary wall with the classic bump-on-a-log appearance as seen in brain pericytes (79, 80). Primary processes from heart capillary pericytes extend from the cell body along the long axis of the underlying capillary tube, and secondary circumferential processes originate from these longitudinal stems and wrap at least partway around the vessel circumference, with an average coverage of about 11% of the abluminal vessel surface (79). Furthermore, cardiac pericytes are distinct from those in the brain by their variety and the fact that they frequently have rogue processes that appear to jump from one capillary to another by means of a large diameter (1 micron or more) appendage. This is called a tunneling microtube (Figure 2a) or TMT to distinguish it from the thinner interpericyte tunneling nanotubes, or IP-TNTs, found in the retina (41, 81, 82). These TMTs tunnel extensively between and around ventricular myocytes but with much larger tubes than TNTs. When a TMT lands on a target capillary, it often runs along that capillary for many tens of microns or more (Figure 2a), in contrast to TNTs, which typically terminate on other pericytes (82). We find that TMTs are particularly common in the epicardium and papillary muscles (Figure 2) and allow a pericyte to contact multiple stretches of capillary along its length. The leaps and branches of the TMTs in heart suggest an important signaling or coordinating aspect to their function that presumably complements pericyte contractile and hemodynamic control functions. An additional and possibly important feature of the TMTs is the direct contact with cardiac ventricular myocytes along their route, which may permit important signaling exchanges (Figure 2a,e). In the retina, ∼30% of pericytes exude TNTs (82), and these are found traversing the parenchyma to connect pericytes of adjacent capillary beds, encountering a complex network of neuroglial processes on their way (Figure 2h). These processes appear to play a prominent role in blood flow regulation through Ca2+-dependent mechanisms (82). Clearly, both TMTs and TNTs represent fertile ground for further studies of pericyte contributions to blood flow control and integration of signaling throughout diverse circulatory beds (Supplemental Box 2).
The transcript profile of heart capillary pericytes is also similar to those of brain thin-strand pericytes, in that these cells robustly express Rgs5 (which encodes the signal-transducing molecule regulator of G protein signaling 5) and the ATP-sensitive K+ (KATP) channel subunits Kcnj8 (Kir6.1) and Abcc9 (SUR2), which can be used as markers and also suggest important functional contributions of these proteins (58–60, 76). Cardiac capillary pericytes also express Acta2 (i.e., the contractile protein α-SMA) and Tagln (transgelin), but to a much lower degree than upstream SMCs and contractile pericytes (60).
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