- Top of page
Summary: Purpose: To correlate glucose (and lactate) results obtained from microdialysate to recent studies suggesting that glucose transporter activity may be significantly altered in seizures.
Methods: We used a fluorometric technique to quantify glucose and lactate levels in microdialysates collected from two to four depth electrodes implanted per patient in the temporal and frontal lobes of a series of four patients. Hour-by-hour and day-to-day changes in brain glucose and lactate levels at the same site were recorded. Additionally we compared regional variations in lactate/glucose ratios around the predicted epileptogenic region.
Results: Lactate/glucose ratios in the range of 1–2:1 were the most commonly seen. When the lactate/glucose ratio was <1:1, we typically observed a relative increase in local glucose concentration (rather than decreased lactate), suggesting increased transport, perhaps without increased glycolysis. In some sites, lactate/glucose ratios of 3:1–15:1 were seen, suggesting that a circumscribed zone of inhibition of tricarboxylic acid cycle activity may have been locally induced. In these dialysates, collected from probes closer to the epileptogenic region, the large increase in lactate/glucose ratios was a result of both increased lactate and reduced glucose levels.
Conclusions: We conclude that regional variations in brain extracellular glucose concentrations may be of greater magnitude than previously believed and become even more accentuated in partial seizure patients. Data from concomitant assays of microdialysate lactate and glucose may aid in understanding cerebral metabolism.
Dynamic fluorodeoxyglucose–positron emission tomography (FDG-PET) estimations of influx, efflux, phosphorylation, and dephosphorylation have been performed in patients with complex partial seizures (1–3). There is no complete agreement in these studies. However, reports of significantly decreased glucose extraction (1), FDG influx (2), and lumped constant (3) in the epileptogenic areas all are observations that would be consistent with an alteration of brain capillary glucose transporter activity. Furthermore, because the total surface area of brain cell membranes is considerably greater than the surface area of the blood–brain barrier (BBB) membranes, effective rate-limitation of glucose transport could occur more efficiently at the brain capillary endothelial membranes, rather than the membranes of cells in the brain parenchyma (4). It is known from animal model studies that endothelial glucose transporter maximal velocity is increased by as much as 35% within minutes of an initial seizure (5). However, long-term changes in glucose transport in response to continued intermittent (complex partial) seizures are not yet fully understood.
Recently a form of infantile seizures was described wherein impaired BBB glucose transport, developmental delay, and acquired microcephaly are seen. This condition has been referred to as the glucose transporter protein syndrome, GTPS (6,7), or the Glut1 deficiency syndrome, Glut1DS (8). The diagnostic feature of the syndrome is an unexplained hypoglycorrachia in the clinical setting of an epileptic encephalopathy (7). Blood glucose levels are normal, but the fact that a low-to-normal CSF lactate together with a persistently low CSF glucose is seen in this syndrome (9) prompted our interest in the study of brain extracellular fluid glucose and lactate in adult complex partial seizures.
Furthermore, it has been shown that extracellular brain-nutrient concentrations can be analyzed in vivo from measurements of brain microdialysate obtained from modified depth electrodes (10,11). During et al. (10) characterized ictal and interictal differences in microdialysate and demonstrated that seizures induced a 90% increase in brain lactate levels within the epileptogenic region. They further determined that the increase in lactate persisted for 60–90 min. Fried et al. (11) showed transient ictal increases in glutamate, aspartate, γ-aminobutyric acid (GABA), and taurine (11). Microdialytic monitoring also has been carried out during a variety of neurosurgical interventions (12,13). In many forms of brain insult, such as bypass and aneurysm treatments (14,15), as well as severe head injury (16,17) and subarachnoid hemorrhage (18,19), alterations in both lactate and glucose were reported.
In the present study, we analyzed dialysates from depth electrode/microdialysis probes implanted in patients with intractable complex partial seizures undergoing evaluation for surgical treatment (11). Analyte recovery from different dialysis probes may vary because of probe differences, as well as brain regional differences. Consequently, we analyzed microdialysate glucose relative to lactate as an internal standard. Our objectives were threefold: first, to analyze brain glucose and lactate levels concomitantly in epilepsy patients undergoing dialysate sampling from multiple probes; second, to define possible diurnal alterations or alternatively to determine an absence of diurnal variability in brain glucose (and lactate) levels within epileptogenic regions and adjacent brain; and finally, we sought to determine whether regional lactate/glucose ratios would be uniform or provide information suggesting pathophysiologic alterations in any areas selected for analysis.
- Top of page
In studies of this type, there is a potential for interprobe variability that may be independent of variability in the actual regional concentration of analytes in the extracellular fluid. Sources of variability outside the extracellular fluid may include small variations in pressure and the kinetics of the diffusion of lactate and glucose in different dialysis sites. Recovery also may be affected by changes in supply, metabolism, and in extracellular fluid (ECF) volume (23). Consequently, we can compare (a) changes in lactate and/or glucose within a given probe across time, and (b) changes in the lactate/glucose ratio, with confidence. But possible variability between different probes makes quantitative comparison of absolute regional lactate and glucose concentrations in dialysate samples more problematic.
Hutchinson et al. (13) studied five patients with head injury or subarachnoid hemorrhage and performed a detailed analysis of probe variability in two identical, adjacently situated microdialysis catheters (of the same length, perfusion rate, and perfusion fluid) over a 15- to 24-h interval. Their analyses of glucose and lactate showed small interprobe differences, ranging from 12 to 25%. Considerably larger differences in regional brain glucose concentrations are seen in patients with complex partial seizures, especially in comparing probes adjacent to the epileptogenic site (30–70%; Table 3). If it is assumed that these regional differences in glucose levels are not due solely to interprobe differences, an alternate conclusion is that glucose levels may be altered in and around epileptogenic areas. This conclusion would be consistent with predictions from previous dynamic FDG-PET analyses (3,24). It also suggests that quantitative electron microscopic immunogold analyses of brain capillary Glut1 glucose transporter expression from seizure resections, showing variable amounts of transporter in different areas have a functional significance (24). The latter study showed that small microvolumes of the brain parenchyma had different capillary Glut1 transporter concentrations and therefore could have markedly different glucose levels. As a consequence, adjacent microvolumes of brain tissue might exhibit highly variable glucose utilization rates, on the basis of capillary supply limitation (24). In the present study, our demonstration of regional differences in glucose/lactate ratios around epileptogenic regions in human brain seems consistent with alterations in glucose metabolism.
The implantation of dialysis probes has been likened to an injury model, in which vasogenic edema is attributed to damage caused by probe insertion (25). Benveniste et al. (26) reported that acutely altered brain glucose concentrations were seen in certain sites. Brain glucose utilization rates also are elevated and did not normalize until 24 h had elapsed in these animal model studies (26). In the human brain, Hutchinson et al. (13) recorded similar findings. They demonstrated that in two adjacent dialysis probes, glucose levels from both probes are characterized by initially high, followed by continually decreasing glucose levels through the first 24 h, and relatively stable and uniform glucose concentrations were recorded for the remainder of the study period. We observed a similar pattern in one of our study patients (patient 4; Fig. 4), in whom probes had been implanted on 1 day, and dialysis commenced at 0500 h the following morning. Decreasing glucose levels were recorded only until about noon on the day after surgery, and during the next 24 h, distinctly different dialysate glucose concentrations (characteristic of each of the sites) were seen in each of the three probes (Fig. 4A). We thus postulate that increased glucose levels attributable to local injury associated with probe insertion were apparently resolved during the second postsurgical day, as previously suggested (13,26).
Sokoloff et al. (27) pioneered the development of methods using the phosphorylation and prolonged tissue retention of tracer levels of deoxyglucose phosphate in measuring cerebral metabolic rates. By using [18F]fluorodeoxyglucose, together with positron emission tomography, this method was soon applied to humans (28,29). Noninvasive measurements of the cerebral metabolic rate by the FDG method require precise estimation of the net clearance of FDG, and the isotope correction factor, termed the lumped constant (30). Several reports suggested that uncertainty about the estimates of the individual transfer coefficients (describing influx, efflux, and phosphorylation rates of FDG) has little effect on the final result (31–33). However, the latter studies are based on normal rather than on pathological conditions. Our present results, showing regions of highly variable glucose/lactate ratios around the epileptogenic site, suggest that equally variable glucose influx and metabolism may characterize regions in and around the epileptogenic focus. Although FDG-PET continues to aid in the identification of epileptogenic sites, the alterations in glucose influx make accurate estimations of the lumped constant problematic even during normal (interictal) periods in patients with complex partial seizures.
The use of lumped constants for estimating brain glucose utilization rates with hexose analogues (such as FDG or 2-deoxyglucose) assumes that brain glucose is uniformly distributed in intra- and extracellular spaces of the brain, without significant regional variations. Furthermore, it has been traditionally assumed that glucose exchange between blood and brain was symmetrical. When glucose influx and efflux were equal, the partition volume for glucose and FDG was shown to be the same (34). But quantitative electron microscopic studies show a distinct 3:1 abluminal/luminal asymmetry in (low glucose transporter-expressing) type B capillary membranes. The asymmetry in high glucose transporter–expressing type A endothelia (1:2 abluminal/luminal membrane Glut1) exists in the opposite direction (24). Localized differences in the proportions of these two configurations of glucose transporter–expressing capillaries could contribute to the different glucose concentrations recorded in different brain regions of the patients examined in the present study (Table 3). This observation seems consistent with the prior dynamic FDG-PET studies of Reutens et al. (3), who observed that changes in the lumped constant were seen in epileptogenic areas of their patients.
In one patient (patient 4), in whom we determined glucose and lactate concentrations over an extended (29-h) period, there was no evidence for a significant diurnal rhythm for any of the measures tested in the amygdalae or orbitofrontal cortex. Although preliminary, these data seem to indicate that within a given region, brain glucose levels remained quite constant throughout the sleep–wake cycle (Fig. 4). In contrast, there is a strong diurnal component to peripheral blood glucose levels in humans (35), and FDG-PET studies suggest diurnal variations in cerebral glucose utilization in several neural structures, including the amygdala (36). Studies in rats also have shown relatively little diurnal variation in brain glucose levels, whereas plasma glucose levels and brain glucose utilization rates exhibit a marked diurnal periodicity (37).
In previous measurements of lactate levels in patients with complex partial seizures, During et al. (10) reported that preictal lactate concentrations (150–250 μM) doubled after a seizure, and gradually returned to normal in ∼2 h. The increase to remarkably high lactate levels that we observed interictally (Fig. 2B) has been previously associated with EEG-demonstrable increases in spiking (10). During et al. (10) implanted a 30-mm dialysis probe, whereas a smaller (10-mm) dialysis membrane was noted in the present study, and differences in our lactate levels (Table 3) can be attributed to the dialysis probes. When electrodes are introduced occipitally through the long axis of the hippocampus, with 30-mm dialysis tubing, it is not possible to sample the amygdala or subregions of the hippocampus exclusively (11). Hutchinson et al. (13) also demonstrated that membrane lengths and perfusion rates also significantly affect analyte recovery. In studies of patients with severe head injuries, microdialysate lactate levels ranging from 500 to 2,000 μM by Zauner et al. (22) and 200 to 3,600 μM by Menzel et al. (16) have been reported, and small (10-mm) microdialysis probes were used in both studies. In minimally traumatized cortex, microdialysate lactate levels from 4-mm probes implanted in brain tissue adjacent to an aneurysm averaged 227 ± 30 μM (±SD, n = 10) (14). Localized brain regions may therefore be characterized by quite extreme changes in both glucose and lactate levels, and we suggest that the rather simple determination of lactate/glucose ratios would permit a more meaningful comparison between different regions as well as different studies.
Human brain lactate and glucose concentrations have been determined from the same microdialysate in a variety of neurosurgical procedures, and technical differences between the different studies tend to be normalized when the ratios of these two analytes are compared. In cases of severe acute head trauma, mean lactate/glucose ratios of ∼1:1 were reported in patients with a good outcome, 1–2:1 in patients with moderate to severe disability, and >2:1 in patients with poor (death/vegetative) outcomes (22). Studies of Menzel et al. (16) similarly indicate mean lactate/glucose ratios of 1–2:1 in their head-injury patients. Of more relevance to the present study, however, are the analyses of minimally damaged cortex carried out by Reinstrap et al. (38), Langemann et al. (39), and Bachli et al. (14). Microdialysate lactate and glucose concentrations were measured before probe removal, in a series of patients in which the neurosurgical intervention was to treat an unruptured aneurysm. Mean lactate/glucose ratios of 0.86 ± 0.11 were recorded (14). It also was shown in one patient that in the 2 h of retraction during surgical treatment of an aneurysm (in the posterior communicating artery), lactate/glucose ratios were consistently >2:1, but rapidly changed to <1:1 in the 3 h after retraction pressure was withdrawn, and the ischemic stress is removed (14). Reinstrap et al. (38) reported baseline values of glucose (1.7 ± 0.9 mM) and lactate (2.9 ± 0.9 mM) in nine patients and saw a lactate/glucose ratio of 1.7. In anesthetized patients, the lactate/glucose ratio of minimally disturbed brain (1.17) was slightly lower (39). Data from our patients seem to suggest that dialysate lactate/glucose ratios fall into one of three separate groups (Table 3). First, lactate/glucose ratios of <1 were occasionally recorded (Fig. 2), and this occurrence seems to be associated with elevated brain glucose levels. Second, lactate/glucose ratios of 1–2:1 were the most frequently recorded, in the present study (Figs. 1–3) and in others (16,22,38,39), suggesting this represents the usual state of glucose use. Third, lactate/glucose ratios of >2 (Figs. 2 and 4) were observed in selected regions, presumably linked with some sort of pathophysiologic event. In epilepsy patients, these markedly elevated lactate/glucose ratios are coincident with elevations in lactate levels (Figs. 2 and 4; Table 2), and increased spiking activity is associated with similar increases in lactate (10). However, in aneurysm patients, the transiently elevated lactate/glucose ratio was associated with an ischemic response (to retraction pressure) and were coincident with a reduction in brain glucose. Some minutes after retraction pressure was eased, dialysate glucose levels increased, and the lactate/glucose ratio normalized (14). The reduction in brain glucose may be attributable to the well-established acute downregulation of BBB glucose transport seen in anoxia, which normalizes after the stress is relieved (40). Ischemia due to retraction pressure also was seen to increase microdialysate lactate/glucose ratios in another study (15). Thus elevated (>2) lactate/glucose ratios occur in brain via different mechanisms. In epilepsy patients, they are attributable to increased lactate accumulation, presumably associated with reduced oxidative activity. In contrast, the transiently elevated lactate/glucose ratios that accompany retraction ischemia apparently are explained by transient downregulation of glucose transport.
In conclusion, regional variations in brain extracellular glucose concentrations may be of greater magnitude than previously established. In two of the four patients with partial seizures whom we examined interictally, regional lactate/glucose ratios differed by log orders of magnitude, and changes in the lactate/glucose ratios are believed to involve alterations in both regional brain glucose and lactate. Changes in brain glucose are presumably attributable to alterations in brain capillary glucose transporter expression described in previous quantitative immunogold electron microscopic studies of seizure resections (24). The elevated regional lactate levels that also were recorded in two of the four patients examined is seemingly consistent with the observation of During et al. (10), who demonstrated that increases in lactate were seen both ictally and in the absence of active seizures. Furthermore, we observed lower glucose concentrations in the right amygdalae (Table 2) in patients with a right (or bilateral) epileptogenic focus (Table 1), and relatively lower glucose metabolism has been seen by others in the (right) hippocampus, ipsilateral to temporal lobe epileptogenic zones (41). An underlying common mechanism may therefore explain our observation of asymmetric glucose levels in left compared with right amygdalae (Table 2), as well as the Asymmetry Index in hippocampal glucose metabolic rate (and hippocampal volume), reported in patients with temporal lobe epilepsy (41). The present study establishes that modified depth electrodes can be adapted to compare dialysate analyte concentrations diurnally, and in several different brain regions simultaneously. Concomitant assays of microdialysate lactate and glucose may augment FDG-PET analyses of cerebral metabolism in seizure disorders, and help identify sites where rates of glucose transport, glycolysis, and oxidative metabolism may differ.
Since the manuscript was accepted for publication, another study has analyzed lactate and glucose concentrations from microdialysate probes located in nonepileptic regions of patients with complex partial seizures (42). When plasma glucose levels were normal (5.5 mmol/L) the dialysate lactate/glucose ratios averaged 1.7 (n = 12). When hyperglycemia was induced (plasma glucose level = 11.5 mmol/L) the microdialysate lactate/glucose ratio was 0.95. However, in hypoglycemic conditions (plasma glucose = 3 mmol/L) microdialysate lactate/glucose ratios averaged 6.1 (42). These observations are consistent with our suggestion that microdialysate lactate/glucose ratios of 1 to 2 would be anticipated under normal conditions, and brain microdialysate lactate/glucose ratios above or below the 1 to 2 range are associated with pathophysiological events.