Glucose
Glucose and O2 are the most important substrates for brain energy metabolism. Glucose enters ISF across the blood–brain barrier via the more glycosylated form of a passive, selective carrier, GLUT1 (SLC2A1), that is present in membranes located on both surfaces of the endothelial cells. From ISF it rapidly enters both astrocytes by the less glycosylated form of GLUT1 and neurons via GLUT3 (see Fig. 11). The rate-limiting step in glucose metabolism is the effectively irreversible phosphorylation by hexokinase. Normally glucose influx into the parenchyma is higher than the rate of phosphorylation, and thus there must be some efflux corresponding to the difference. This efflux is also primarily across the blood–brain barrier via GLUT1. Because both influx and efflux of glucose take place by passive transport there is no additional metabolic cost caused by having influx greater than the metabolic rate.
Fig. 11 GLUT and MCT transporters at or near the blood–brain barrier. GLUT1 and MCT1 are present on endothelial cells; GLUT1 and MCT4 on astrocytes
(Figure simplified and redrawn from Simpson et al. [315])
It has long been known that glucose is able to cross the blood–brain barrier rapidly [189, 299–302]. Crone [299] found that at low concentrations as much as 50% of the glucose arriving in the arterial blood could be extracted in a single pass, but that this percentage decreased with concentration, falling to 28% at 5 mM and ~ 14% at 14 mM. This extensive but saturable transport implies the presence of a specific transporter, which as stated above is GLUT1 (SLC2A1) [303–305].
The expression of GLUT1 in the endothelial cell membranes has been measured in several different ways: by cytochalasin-B binding, by specific antibody binding, and by proteomic methods (see Table 2 for references). In the proteomic studies from the group of Terasaki, Uchida, Ohtsuki and colleagues, GLUT1 was found to be the most highly expressed of all the transporters that are present in the membranes of the endothelial cells [306].
Table 2 Expression of GLUT1 at the blood–brain barrier
Source Methoda Species Expression/pmol mg−1
Luminal Totalb Abluminal
Relative to microvessel protein
Dick et al. [303] Cytochalasin B binding Rat, pig 69–80
Kalaria et al. [571] Cytochalasin B binding Human 42
Farrell and Pardridge [572] Immunogold e.m Rat 12%c 100%c 48%c
Cornford et al. [573] Immunogold e.m Human 48% 100% 18%
Vannucci et al. [574] Cytochalasin B binding Rat 40–125
Kamiie et al. [182] Proteomics Mouse 90
Uchida et al. [184] Proteomics Human 139
Shawahna et al. [176] Proteomics Human 78.5
Hoshi et al. [185] Lysate digestion proteomics Rat 84–98
Relative to membrane protein
Simpson et al. [575] Fractionation, cytochalasin B binding Bovine 620 280
Kubo et al. [58] Fractionation, proteomics Porcine 79% 21%
Zhang et al. [188] Proteomics Porcine 300
aFractionation = fractionation of isolated plasma membranes
bFor immunogold detection, values are percentages of the immunogold particles where the total includes cytoplasmic
cAntigen for the antibody used by Farrell and Pardridge appears to be partially masked for GLUT1 most markedly in the luminal membrane in bovine endothelial cells [575]
A rough estimate of the glucose clearance in man can be calculated from the rate of consumption, about 0.55 mol day−1 = 380 µmol min−1 [295, 296] or, for a 1400 g brain, 270 nmol g−1 min−1. For a difference between the concentrations in plasma and ISF of 5 mM this corresponds to CL ~ 54 µL g−1 min−1. In isolated perfused dog brains Betz et al. [302] measured the loss of glucose from the blood flow through the brain and found about 0.6 µmol g−1 min−1 at 6 mM from which at this concentration CL = 100 µL g−1 min−1. Hawkins [307] lists values ranging from 158 to 352 µL g−1 min−1 (at 6 mM glucose) depending on brain region (inferior colliculus the highest). Note that the first two of the estimates above are based on the net flux of glucose while the values listed by Hawkins are based on the unidirectional influx. Because all of these estimates far exceed the clearance expected for perivascular efflux, ~ 1 µL g−1 min−1 (see Sect. 3 and Table 1), the perivascular route is likely to be of minor importance.
Cutler and Sipe [301] using anaesthetized cats, Bachelard et al. [308, 309] using anaesthetized rats and Betz et al. [302] using isolated perfused dog brains all found that the influx of glucose measured using tracers could exceed the net flux by two to threefold. This is a direct, experimental demonstration that there is efflux across the blood–brain barrier that can be as large as two-thirds of the influx. This would of course be less under conditions of increased metabolic demand.
Glucose distributes rapidly between intracellular and extracellular water within the parenchyma and thus its volume of distribution is close to the total aqueous volume, which is VD = 0. 77 mL g−1 [310–315].14 Pfeuffer et al. [316] used diffusion weighted NMR to distinguish between intracellular and extracellular glucose and found that only 19% of the glucose in the parenchyma was extracellular which is in agreement with the fraction of water that is extracellular. These observations imply that glucose transport across the membranes of astrocytes and neurons is rapid compared to the rate of metabolism.
When glucose concentrations in plasma are near 6 mM, the average concentration of glucose in brain water is roughly 1.3 mM (see Sect. 5.3.2). Even two fold changes in the concentration in brain water have little effect on the cerebral metabolic rate of glucose, CMRglc, because these concentrations are substantially greater than the Km for phosphorylation of glucose by hexokinase (0.04–0.05 mM [317–319]) and hence hexokinase, the first step in glucose metabolism, remains nearly saturated (compare e.g. [313]).
It is unclear why the passive glucose transport at the blood–brain barrier is mediated by a carrier rather than by a pore. Pores have the advantage that they do not undergo any large conformation changes during transport of each substrate. Hence they are capable of high turnover numbers, which would seem to be an advantage. On the other hand carriers allow more complicated coupling of transport between different solutes and it is possible that during transport of a relative large solute like glucose, it is easier for a carrier than the “open hole” of a pore to prevent unwanted transfer of other solutes. (Water can probably get through both carriers and pores. The possibility that water permeability of GLUT1 may or may not be important at the blood–brain barrier [320] was considered in Section 4.3.6, footnote 17 of [4]). While arguments for “why a carrier” are speculative, the structural and kinetic evidence, reviewed in the following subsections, leave little doubt but that glucose transport across the membranes of the endothelial cells of the blood–brain barrier is mediated by a carrier.

Structure of GLUT1 (SLC2A1) and the kinetics of the glucose transport it mediates in red blood cells
A crystal structure for GLUT1 has been obtained using a GLUT1 construct purified from an expression system (see Fig. 12) [321]. In this structure a bundle of α-helices spans the membrane surrounding an inner cavity open at the cytoplasmic end. This structure and those for related transporters (for references see [322]) strongly support the widely held view that the transport kinetics should be described using a carrier model (see Appendix D). A binding site in the central cavity of the carrier can be exposed to either side of the membrane, but only one side at a time. While the site is exposed a substrate molecule can associate with or dissociate from the site. The side of exposure can be altered by a conformation change in the carrier and the substrate can then associate or dissociate on the other side of the membrane.
Fig. 12 Structure of the human glucose transporter GLUT1. The structure of full-length human GLUT1 containing two point mutations (N45T, E329Q) was determined in an inward-open conformation. The side and cytoplasmic views are shown. The corresponding transmembrane segments in the four 3-helix repeats are coloured the same. The extracellular and intracellular helices are coloured blue and orange, respectively. A slab of cut- open view of the surface electrostatic potential, which was calculated with PyMol50, is shown on the right to facilitate visualization of the inward-facing cavity. IC indicates intracellular helix. Reprinted by permission from Springer Nature from Nature 510, 121–126, Crystal structure of the human glucose transporter GLUT1 by Deng et al. [321]
Since GLUT1 is highly expressed in red blood cells, they have been used as the most convenient system in which to study the kinetics of its transport. There are two prominent features revealed by these studies that must be accommodated in any model. On the one hand the normal net transport of glucose occurs without input of energy from any source other than the concentration gradient, on the other hand downhill movement of one type of sugar can be coupled to uphill movement of another (see Fig. 13), a phenomenon called counter-flow or counter-transport [322–325]. A closely related phenomenon is trans-stimulation, an increase in influx when internal concentration is increased or an increase in efflux when external concentration is increased (see Fig. 13 and, for a quantitative example, Appendix D). In terms of a simple carrier model, the observation of net glucose transport when it is the only substrate implies that both the loaded and unloaded forms of the carrier can change conformation thus altering exposure of the binding site. This allows transport of solute in one direction to occur without transport in the opposite direction, i.e. the transport is not an obligatory exchange. Similarly counter-transport or trans-stimulation imply that the rate constants for the conformation changes when the carrier is loaded are at least comparable to those for the unloaded carrier so that solute on the trans side can assist transport from the cis side by increasing the rate of return of the carrier.
Fig. 13 Interpretation of net flux of a single solute, obligatory exchange, and trans-stimulation in terms of a simple carrier model. In each case the concentration of the first solute (filled black circle) is higher on the cis side (left) than on the trans side (right). a Net flux of solute from cis to trans is supported by return of the free carrier. b If return of the carrier is only possible with a solute bound, there is obligatory exchange, either self-exchange or counter-transport of another solute (circle). c Trans-stimulation is a combination of these two effects. Flux of the first solute from cis to trans can be increased if there is more solute (either sort) on the trans side (here the right) provided that increases the rate of return of the carrier—i.e. it increases the rate of conformation changes of the carrier from trans-facing to cis-facing
Trans-stimulation can markedly increase influx and efflux of glucose at high glucose concentrations (see Appendix D) and it is therefore very important in studies of the mechanism of transport. However, it has little if any effect on the net flux and it is the net flux that is important for the delivery of glucose for metabolism. The exchanges underlying trans-stimulation are likely to be much more important for large neutral amino acids where several compete for transport by the same carrier (see Sect. 5.5).
The kinetics of the simple carrier model are complex even in the steady-state [325–329]. GLUT1 (SLC2A1) kinetics are complicated further by the added twist that the GLUT1 protein may exist in the membranes as part of a homo-tetramer, each capable of transport, but in a coupled manner such that transport through one affects the transport through the others [322, 330]. Given these and further complexities considered in the next section, it should not be surprising that definitive characterization of glucose transport at the blood–brain barrier remains elusive (see Appendix D).

Glucose transport kinetics at the blood–brain barrier
Transport of glucose into and out of the brain is clearly more complex than that into and out of red blood cells. Firstly GLUT1 is needed in both membranes of the endothelial cells of the blood–brain barrier to allow the glucose to enter on one side and leave on the other. However, because the endothelial cells are very thin and correspondingly contain very little glucose, provided that the properties of the transport in the two membranes are similar, it is thought that the transport can still be described, at least qualitatively, as transport across a single barrier [331–333]. Secondly once across the blood–brain barrier, glucose is metabolized at a rate comparable to the rates of influx and efflux across the barrier while in red blood cells transport is much faster than metabolism. Thirdly there is also the technical difficulty that, with the important exception of the study in 1975 by Betz et al. [327], it has not proved possible to manipulate interstitial fluid glucose concentrations during the experiments. In most studies all that has been done is either to measure the extraction of glucose (total or labelled) from blood as described above or to measure the variation in the total amount of glucose present in the parenchyma with time as a function of glucose concentration in plasma. Mason et al. [334] compare the results obtained in many studies performed prior to 1992 but with the surprising omission of reference to studies from Betz’s group. Also in 1992, Gjedde [335] reviewed results obtained for glucose transport in rat and man. Glucose transport into and within the brain has been analyzed and reviewed by Simpson et al. [315], Barros et al. [314] and, more recently, by Patching [336].
In one of the first attempts to establish the mechanism of glucose transport at the blood–brain barrier, Buschiazzo et al. [319] found that 3-O-methyl-d-glucose, a non-metabolizable derivative of glucose, competes with glucose for transport, and furthermore that an inward gradient of glucose could drive 3-O-methyl-d-glucose uphill out of the brain, i.e. there is counter-transport for GLUT1 at the blood–brain barrier just as in red blood cells. Further evidence that GLUT1 behaves in a similar manner in the two environments was obtained by Betz et al. [327] who found that the rate of glucose influx was increased by increasing the concentration of glucose within the brain, i.e. there is trans-stimulation (see Appendix D).
Buschiazzo et al. [319] and Betz et al. [327] determined the total glucose in the parenchyma for different glucose concentrations in plasma (see Fig. 14). Subsequently NMR has been used to measure glucose content in conscious humans and lightly anaesthetized rats [334, 337–341]. The NMR results for humans and rats confirm under nearly physiological conditions (see Fig. 14) that brain glucose content continues to increase with plasma concentration for plasma concentrations up to at least 30 mM well above a typical resting value, 6 mM. They also confirm that the rates of glucose influx and efflux are respectively larger than and not much smaller than the rate of metabolism. Because influx and efflux substantially exceed the expected efflux via the perivascular route, the net flux across the blood–brain barrier is normally taken to be equal to CMRglc at steady-state.
Fig. 14 Four studies of brain glucose content versus glucose concentration in blood. In two studies glucose content was measured by chemical assay, a in anaesthetized rats by Buschiazzo et al. [319] and b in isolated perfused brains from dogs by Betz et al. [327]. In the latter it was assumed that brain water was 0.75 mL g−1. In the other two studies glucose content was determined by magnetic resonance spectroscopy, c in conscious humans by Gruetter et al. [337] and d in lightly anaesthetized rats by Choi et al. [338]. In all studies the glucose content continues to increase with plasma concentration even though it is known that the influx of glucose shows saturation. The explanation is that efflux also saturates and the increase in content must parallel the increase in plasma concentration in order for efflux to increase so that it is equal to influx minus the constant rate of glucose metabolism (see Appendix D)
In the results reported by Duarte et al. (see Figure 3 in [341]) using rats, following a step change in cplasma from 4 to 20 mM the brain content of glucose increased from about 0.5 to 4.5 µmol g−1 with a half life of about 16 min which indicates a net rate of accumulation of 0.122 µmol g−1 min−1, i.e. using their value of CMRglc, 0.52 µmol g−1 min−1, there is an influx of 0.64 µmol g−1 min−1 which is similar to that reported by Betz et al. in 1974 [302] for the dog.
It has so far not proved possible to analyse glucose efflux directly after injection of glucose into the brain. Any such measurements face major challenges including separating efflux from metabolism and avoiding disturbance of the efflux processes by the injection or infusion. The study by Ball et al. [85] established that during a 5 min, 0.1 µL min−1 infusion into the inferior colliculus glucose can move, presumably by a perivascular route, to the adjacent meninges strongly suggesting that as expected there is perivascular efflux of glucose. However, estimating the normal rate of this process to see if the perivascular clearance notably exceeds the 1 µL g−1 min−1 found in other regions would require measurement of the time course of the appearance of glucose in the meninges after the end of the infusion.15
The glucose efflux across the blood–brain barrier can be calculated if the influx and net flux are both known as indicated earlier in this discussion of glucose. Furthermore if it can be assumed that the fluxes are described by the expressions of the form derived from the carrier model, the rate of efflux can be calculated from the measured rates of influx versus the concentrations in plasma and ISF. An example of this using the data from Betz et al. [327] for the isolated perfused dog brain is given in Appendix D and Additional file 1. These data remain the only measurements of glucose influx versus plasma concentration for a range of known concentrations within the brain. Hence the calculated results in Appendix D are the only available results for efflux as a function of both plasma and ISF concentrations.
The fits to the data of Betz et al. [327] (see Additional file 1) indicate that the net flux = CMRglc for cplasma = 6 mM is 0.65 µmol g−1 min−1 with cisf = 1.2 mM. This value of CMRglc is close to those expected for rats but about twice that for humans. The fits also predict that glucose consumption, CMRglc, could increase to about 0.9 µmol g−1 min−1 with cisf approaching 0 without any change in transport capacity. However, larger increases in glucose consumption are required in order to support nervous activity. Changes in transport capacity are considered in Sect. 6.2.
Both neurons and astrocytes have transporters that will allow uptake of glucose and both can use it as a substrate for energy production. The proportions of glucose metabolism that occur in astrocytes and neurons remain controversial [315, 342–346] (see next section).