Local Glucose Concentration Research Articles

Glucose sensing by hypothalamic neurones and pancreatic islet cells: AMPle evidence for common mechanisms?

A fuller understanding of the central mechanisms involved in controlling food intake and metabolism is likely to be crucial for developing treatments to combat the growing problem of obesity in Westernised societies. Within the hypothalamus, specialized neurones respond to both appetite-regulating hormones and circulating metabolites to regulate feeding behaviour accordingly. Thus, the activity of hypothalamic glucose-excited and glucose-inhibited neurones is increased or decreased, respectively, by an increase in local glucose concentration. These 'glucose-sensing' neurones may therefore play a key role in the central regulation of food intake and potentially in the regulation of blood glucose concentrations. Whilst the intracellular signalling mechanisms through which glucose-sensing neurones detect changes in the concentration of the sugar have been investigated quite extensively, many elements remain poorly understood. Furthermore, the similarities, or otherwise, with other nutrient-sensing cells, including pancreatic islet cells, are not completely resolved. In this review, we discuss recent advances in this field and explore the potential involvement of AMP-activated protein kinase and other nutrient-regulated protein kinases.

Continuous Glucose Monitoring by Means of Fiber-Based, Mid-Infrared Laser Spectroscopy

The application of mid-infrared laser spectroscopy to the reagent-free quantification of the concentration of glucose was investigated using cryogenically cooled lead salt lasers or, alternatively, quantum cascade lasers operating at room temperature. The concentration of glucose in aqueous solutions was quantified by means of fiber-based attenuated total reflection (ATR) spectroscopy (fiber-optical evanescent field analysis, FEFA) as well as fiber-based transmission spectroscopy. Both methods have the potential to be utilized by small fiber sensors, which can be inserted into the subcutaneous tissue in order to continuously measure the local concentration of glucose. In our in vitro experiments, noise-equivalent concentrations as low as 10 mg/dL were achieved. The mid-term stability of the measurement schemes was investigated by means of Allan variance analysis. Based on the research presented in this manuscript, an all-room-temperature measurement scheme using quantum cascade lasers, miniaturized fiber-optic sensors, and pyroelectric detectors appears well suited for the continuous monitoring of glucose concentrations at physiological levels.

Modeling the dynamics of E. coli populations in the three-dimensional turbulent field of a stirred-tank bioreactor—A structured–segregated approach

In the present work, an Euler–Lagrange approach has been applied to characterize the behavior of a heterogeneous cell population in a stirred-tank bioreactor with non-ideal mixing. It allows one to describe population behavior as the outcome of the interaction between the intracellular state of its individual cell and the turbulent flow field in the reactor. The modeling approach and the numerical method employed are based on an Euler–Lagrange formulation of the system combined with a fractional-step method to allow for a stable, accurate, and numerically efficient solution of the underlying equations. This strategy permits one to account for the heterogeneity present in real reactors in both the abiotic and biotic phases. The example of sugar uptake (phosphotransferase system, PTS) of E. coli cells growing in a fed-batch culture is used to illustrate the application of the approach. The activity of the uptake system depends on the local concentration of glucose as well as the ratio of the intracellular concentrations of phosphoenolpyruvate and pyruvate, which in turn is a function of the history of the individual cell. The simulation results point to distinct differences in the viability of the cells at different scale of operation.

Glucose-responsive vesicular sensor based on boronic acid–glucose recognition in the ARS/PBA/DBBTAB covesicles

The amphiphile, 4-(4-dodecyloxybiphenyl-4-yloxy) butyl trimethylammonium bromide (DBBTAB), has a cationic group with quaternary ammonium salt, which can self-organize into vesicles in dilute aqueous solutions. The alizarin red S (ARS) with a negatively charged group is attracted electrostatically to the positively charged surface on the DBBTAB vesicles to form covesicles. By taking advantages of the features, a vesicular fluorescent sensor was prepared based on phenylboronic acid (PBA)–glucose recognition in the aqueous ARS/PBA/DBBTAB covesicles. The sensor was constructed with three constituents: PBA, DBBTAB amphiphile and ARS which served as the detector. The vesicular sensor enhanced the sensitivity to glucose by about seven- to eight-fold, compared with the same aqueous PBA/ARS solution, which was ascribed to the increased local concentrations of glucose on the surface of vesicles. The vesicular sensor may be available for detection of saccharides in biological systems.

The metabolism of the diabetic foot: in vivo investigation with microdialysis.

Many amputations might be delayed or prevented by more effective clinical supervision of the diabetic foot ulcer. The aim of this study was to measure the local metabolism on the edge of a diabetic ulcer and compare it to healthy subcutaneous tissue. In 5 non-fasting diabetic patients, we inserted a microdialysis catheter into the edge of a diabetic ulcer and a reference catheter into healthy abdominal subcutaneous tissue, and recorded the local concentrations of glucose, lactate and glycerol during rest. The concentrations of glucose in the ulcers were 7.8 mM (SEM 1.9) and in the reference tissue 10.6 mM (SEM 1.8) (p = 0.4). The concentrations of lactate were 2.9 mM (SEM 0.7) and 2.1 mM (SEM 0.7) (p = 0.2), while those of glycerol were 290 microM (SEM 84) vs 98 microM (SEM 7.2) (p = 0.002). This study shows that microdialysis can detect differences in dialysate concentrations of metabolites in diabetic ulcers and a reference tissue, providing valuable information concerning metabolites in the diabetic foot ulcer. Future studies should combine the technique with measurements of local blood flow.

Regional analyses of CNS microdialysate glucose and lactate in seizure patients.

To correlate glucose (and lactate) results obtained from microdialysate to recent studies suggesting that glucose transporter activity may be significantly altered in seizures. 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. 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. 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.

Integration of bienzymatic disaccharide sensors for simultaneous determination of disaccharides by means of light addressable potentiometric sensor

Three saccharides such as sucrose, maltose and glucose were simultaneously determined by the use of the light addressable potentiometric sensor (LAPS). One-chip integration of three saccharide sensors was performed by means of a glucose sensor using glucokinase. By the addition of oligosaccharide hydrolases such as invertase and α- d-glucosidase to the glucose sensors, the disaccharide concentrations were determined by variation of glucose concentrations. All the enzymes employed were thermostable enzymes because of their durability for repeated measurements. Clear distinctions of the response patterns were obtained among the three saccharide samples. Since the simplified LAPS system was limited in number of sensing spots, the complicated chemometrics such as partial least squares (PLS) was not always indispensable. Therefore, a simpler method such as multiple linear regression (MLR) was able to be applied to the mixture analysis under a low-concentrated sample in order to simplify the analysis. Thus, we treated the glucose as a key product in the regression equation optimized for the bienzymatic biosensensing process. The multivariate calibrations of the samples were simultaneously illustrated by using the estimated local glucose concentration. Finally, more than 0.7 of correlation coefficients were obtained between the true and the predicted sample concentrations although the linear approximation was not able to correct a few kinds of inhibition.

GLUT2 in pancreatic and extra-pancreatic gluco-detection (review).

Detection of variations in blood glucose concentrations by pancreatic beta-cells and a subsequent appropriate secretion of insulin are key events in the control of glucose homeostasis. Because a decreased capability to sense glycemic changes is a hallmark of type 2 diabetes, the glucose signalling pathway leading to insulin secretion in pancreatic beta-cells has been extensively studied. This signalling mechanism depends on glucose metabolism and requires the presence of specific molecules such as GLUT2, glucokinase and the K(ATP) channel subunits Kir6.2 and SUR1. Other cells are also able to sense variations in glycemia or in local glucose concentrations and to modulate different physiological functions participating in the general control of glucose and energy homeostasis. These include cells forming the hepatoportal vein glucose sensor, which controls glucose storage in the liver, counterregulation, food intake and glucose utilization by peripheral tissues and neurons in the hypothalamus and brainstem whose firing rates are modulated by local variations in glucose concentrations or, when not protected by a blood-brain barrier, directly by changes in blood glucose levels. These glucose-sensing neurons are involved in the control of insulin and glucagon secretion, food intake and energy expenditure. Here, recent physiological studies performed with GLUT2-/- mice will be described, which indicate that this transporter is essential for glucose sensing by pancreatic beta-cells, by the hepatoportal sensor and by sensors, probably located centrally, which control activity of the autonomic nervous system and stimulate glucagon secretion. These studies may pave the way to a fine dissection of the molecular and cellular components of extra-pancreatic glucose sensors involved in the control of glucose and energy homeostasis.

Paracellular glucose transport plays a minor role in the unanesthetized dog.

Traditionally, intestinal glucose absorption was thought to occur through active, carrier-mediated transport. However, proponents of paracellular transport have argued that previous experiments neglected effects of solvent drag coming from high local concentrations of glucose at the brush-border membrane. The purpose of this study was to evaluate glucose absorption in the awake dog under conditions that would maximize any contribution of paracellular transport. Jejunal Thiry-Vella loops were constructed in six female mongrel dogs. After surgical recovery, isotonic buffers containing L-glucose as the probe for paracellular permeability were given over 2-h periods by constant infusion pump. At physiological concentrations of D-glucose (1-50 mM), the fractional absorption of L-glucose was only 4-7% of total glucose absorption. Infusion of supraphysiological concentrations (150 mM) of D-glucose, D-maltose, or D-mannitol yielded low-fractional absorptions of L-glucose (2-5%), so too did complex or nonabsorbable carbohydrates. In all experiments, there was significant fractional water absorption (5-19%), a prerequisite for solvent drag. Therefore, with even up to high concentrations of luminal carbohydrates in the presence of significant water absorption, the relative contribution of paracellular glucose absorption remained low.

Glucose-induced intracellular ion changes in sugar-sensitive hypothalamic neurons.

In the lateral hypothalamic area (LHA) of rat brain, approximately 30% of cells showed sensitivity to small changes in local concentrations of glucose. These "glucose-sensitive" neurons demonstrated four types of behavior, three of which probably represent segments of a continuous spectrum of recruitment in response to ever more severe changes in blood sugar. Type I cells showed maximum activity </=5.6 mM blood glucose but became completely silent at hyperglycemia of 10-12 mM (normoglycemia 7.6 +/- 0.3 mM; mean +/- SD). Type II and III neurons exhibited a wider range of response. Type IV cells (5-7% of glucose-sensitive neurons) paralleled the behavior of sugar-sensitive cells in ventromedial hypothalamic nucleus (VMH). In VMH, approximately 40% of cells responded to changes in blood glucose over a range of concentrations from 3.6 to 17 mM, by increasing their firing rate as sugar level rose and vice versa. Ionic shifts during increases in blood (brain) glucose levels were similar in LHA types I-III but fastest in I and slowest in III. [Na+]i fell by 5-9 mM, [K+]i rose by 6-8 mM, and plasma membrane hyperpolarized by 5 mV. [Ca2+]i declined by 15-20 nM in line with membrane hyperpolarization. In VMH and type IV LHA cells, [K+]i fell 3-8 mM and plasma membrane depolarized -3 to -5 mV as blood/brain glucose concentration increased from 7.6/2.4 to 17.6/4.2 mM, whereas [Ca2+]i increased from 125 to 180 nM as a consequence of falling membrane potential. During falls in blood/brain sugar concentration the effects in both VMH and LHA cells were reversed. The findings are consistent with the ionic shifts in types I-III LHA cells being dependent on alterations in Na/K-ATPase activity, whereas those in VMH and type IV LHA cells could be caused by modulation of ATP-dependent K+ channels. A possible mechanism for linking the effects of small changes in glucose to ATP generation, which could bring about the above phenomena, is the interposition of a "glucokinase-type" enzyme in a role similar to that which it has in glucose-sensing pancreatic beta-cells.

Effect of Ischemia and Reperfusion on λ of the Lumped Constant of the [14C]Deoxyglucose Technique

We measured the parameter lambda, which is the ratio of the distribution spaces of 2-deoxy-D-glucose (DG) and glucose in the brain, in a model of focal cerebral ischemia in the cat. lambda is the parameter in the lumped constant of the [14C]DG technique most susceptible to changes in ischemia. Cats were subjected to occlusion of the middle cerebral artery for a period of 2 h. During the last 60 min of occlusion, [14C]DG was infused in a programmed fashion so as to obtain a stable arterial blood [14C]DG concentration. The brain was funnel-frozen to preserve tissue metabolites and the frozen brain was sampled regionally (4 to 7-mg samples) for local concentrations of glucose, ATP, phosphocreatine (PCr), and lactate. In a separate series of cats, the infusion of [14C]DG was started after 2 h of occlusion and 3 h of recirculation. In both series, lambda declined slightly for increased levels of tissue glucose and increased appreciably as tissue glucose decreased. A similar relationship was observed between lambda and ATP and PCr, although the correlation was not as clear. Since lambda, and hence the lumped constant, increases in ischemia as well as in postischemic tissue, it is important to measure tissue glucose concentration if quantitative values of local cerebral glucose metabolism are desired in this condition.

Pd—PdO pH microprobe for local pH measurement

Microprobes which can measure local environmental conditions as well as key substrate and product concentrations of fermentation are useful for studying metabolic behaviors of immobilized cells. Oxygen microprobes have been used for such applications. Recently, a fast response glucose microprobe capable of measuring local glucose concentrations in microbial cell layers has been developed. In this communication, we describe a new microprobe suitable for measuring local pH in immobilized cell layers

Glucose modulation of somatostatin and LHRH release from rat hypothalamic fragments in vitro.

In the rat, hypoglycaemia inhibits growth hormone secretion, but the mechanism is unclear. To investigate this further, we have studied the effects of glucose and 2-deoxy-D-glucose on somatostatin and LHRH release from rat hypothalamic fragments incubated in vitro. Glucose (1.35-22 mM) was added to glucose-free medium and 5 and 50 mM 2-deoxy-D-glucose were added to medium containing 5.5 mM glucose. Medium somatostatin and LHRH levels were measured by RIA. Somatostatin and LHRH released diluted in parallel with synthetic somatostatin and LHRH. Sephadex gel filtration demonstrated two molecular forms of somatostatin, 70% coeluting with somatostatin-14 and 30% with somatostatin-28; LHRH coeluted with synthetic LHRH. KCl (30-100 mM) resulted in release of somatostatin and LHRH; this was reduced in calcium-free medium. Basal and K+-stimulated somatostatin release were significantly increased by reducing glucose levels (r = -0.6, p less than 0.001). Basal LHRH was not influenced by glucose. Basal and K+-induced somatostatin release were significantly increased by 2-deoxy-D-glucose (p less than 0.05), while LHRH levels remained unchanged. Our results demonstrate that basal and K+-induced somatostatin release from rat hypothalamic fragments are modulated by local glucose concentrations, and this effect is specific as it is not paralleled by LHRH changes. We suggest that the reduction in growth hormone secretion during hypoglycaemia in the rat might be mediated, at least in part, via a direct effect of glucose on somatostatin release.