Anders Ekholm

Extra- and Intracellular pH in the Brain During Ischaemia, Related to Tissue Lactate Content in Normo- and Hypercapnic rats.

The objective of the present study was to assess the relationship between the amount of lactate accumulated during complete ischaemia and the ensuing changes in extra‐ and intracellular pH (pHe and pHi respectively). The preischaemic plasma glucose concentration of anaesthetized rats was varied by administration of glucose or insulin, pHe was determined in neocortex with ion‐sensitive microelectrodes, and tissue lactate and CO2 contents were measured, tissue CO2 tension being known from separate experiments. The experiments were carried out in both normocapnic [arterial CO2 tension (PaCO2) ‐40 mm Hg] and hypercapnic (PaCO2 ‐80 mm Hg) animals. Irrespective of the preischaemic CO2 tension, δpHe was linearly related to tissue lactate content. Depending on the preischaemic glucose concentration, δpHe varied from <0.4 to >1.4 units. The results thus fail to confirm previous results that the changes in pHe describe two plateau functions (δ pHe‐0.5 and 1.1, respectively), with a transition zone at tissue lactate contents of 17–20 mmol kg−1. Changes in pH; given in this study are based on the assumption of a uniform intracellular space. The pH, changed from a normal value of ‐7.0 to 6.5, 6.1 and 5.8 at tissue lactate contents of 10, 20 and 30 mmol kg‐1. The intrinsic (non‐bicarbonate) buffer capacity, derived from these figures, was 23 mmol kg −1 pH−1. Some differences in pH and in HCO3− concentration between extra‐ and intracellular fluids persisted in the ischaemic tissue. These differences were probably caused by a persisting membrane potential in the ischaemic cells.

Phosphorylase a and Labile Metabolites During Anoxia: Correlation to Membrane Fluxes of K+ and Ca2+

Abstract: The objective of the present study was to explore mechanisms responsible for activation of ion conductances in the initial phases of brain ischemia, particularly for the early release of K+ that precedes massive cell depolarization, and rapid downhill fluxes of K+, Na+, Cl−, and Ca2+. As it has been speculated that a K+ conductance can be activated either by an increase in the free cytosolic calcium concentration (Ca2+i) or by a fall in ATP concentration, the question arises whether the early increase in extracellular K+ concentration (K+e) is preceded by a rise in Ca2+i and/or a fall in ATP content. In the present experiments, ischemia was induced in rats by cardiac arrest, the time courses of the rise in K+e and cellular depolarization were determined by microelectrodes, and the tissue was frozen in situ through the exposed dura for measurements of levels of labile metabolites, including adenine nucleotides and cyclic AMP (cAMP), after ischemic periods of 15, 30, 60, and 120 s. Conversion of phosphorylase b to a was assessed, because it depends, among other things, on changes in Ca2+i. The K+e value rose within a few seconds following induction of ischemia, but massive depolarization (which is accompanied by influx of calcium) did not occur until after ∼65 s. Activation of phosphorylase was observed already after 15 s and before glycogenolysis had begun. At that time, 3′,5′‐cAMP concentrations were unchanged, and total 5′‐AMP concentrations were only moderately increased. The results demonstrate that a K+ conductance is activated at a time when the overall ATP concentration remains at 95% of control values. If major compartmentation can be excluded, the results fail to demonstrate that an ATP‐activated K+ conductance is involved. In view of the early activation of phosphorylase, one may speculate that the triggering event is a rise in Ca2+i.

Extracellular pH in the brain during ischemia: relationship to the severity of lactic acidosis.

On the basis of data showing a bimodal distribution of values for extracellular pH (pHe), and a discontinuous ΔPco2/Δlactate relationship, Kraig et al. (1986) proposed that H+ is grossly compartmentalized between neurons and glia in the ischemic brain. We measured ΔpHe during ischemia, varying ischemic lactate contents between 9 and 38 mmol kg−1. No bimodal distribution was found, but ΔpHe varied linearly with lactate content. Because we have also failed to record a discontinuous ΔPco2/Δlactate relationship, we conclude that major compartmentation of H+ does not occur during ischemia.

Changes of labile metabolites during anoxia in moderately hypo- and hyperthermic rats: correlation to membrane fluxes of K+.

The objective of this study was to assess the influence of temperature on the coupling among energy failure, depolarization, and ionic fluxes during anoxia. To that end, we induced anoxia by cardiac arrest in anesthetized rats maintained at a body temperature of either 34 degrees C or 40 degrees C, measured extracellular K+ concentration (K+e), and froze the neocortex through the exposed dura for measurements of phosphocreatine (PCr), creatine (Cr), ATP, ADP, and AMP, glucose, glycogen, pyruvate and lactate content after ischemic intervals of maximally 130 s. Free ADP (ADPf) concentrations were derived from the creatine kinase equilibrium. Hypothermia reduced the initial rate of rise in K+e, and delayed the terminal depolarization; however, both hypo- and hyperthermic animals showed massive loss of ion homeostasis at a K+e of 10-15 mM. The initial rate of rise in K+e did not correlate to changes in ATP, or ATP/ADPf ratio, suggesting that temperature changes per se may control the degree of activation of K+ conductances. The results clearly showed that, in both hyper- and hypothermic subjects, energy failure preceded the sudden activation of membrane conductances for ions. The results indicate that temperature primarily influences membrane permeability to ions like K+e (and Na+), and that cerebral energy state is secondarily affected. It is proposed that the higher rate of rise of K+e at high temperatures accelerates ATP hydrolysis primarily by enhancing metabolic rate in glial cells.

Chapter 3 Acidosis-related brain damage

Publisher Summary This chapter updates information on the coupling among hyperglycemia, intra and extracellular acidosis and brain damage because of ischemia or hypoxia, and discusses cellular and molecular mechanisms that may be involved. When ischemia is complicated by excessive acidosis, the ischemic damage encompasses post-ischemic seizures, edema, and pannecrosis. The cellular and molecular mechanisms responsible for these alterations have not been adequately defined. However, it seems likely that the acidosis causes damage to inhibitory GABAergic cells by raising Ca i 2+ to levels, which will overload the buffering systems and cause cell death, thereby explaining the proclivity to post-ischemic seizure discharge. The rapidly evolving damage following long periods of ischemia is probably caused by several adverse effects of a raised H + activity: inhibition of Na + /H + exchange and lactate – oxidation, inhibition of mitochondria1 respiration, and acceleration of coupled Na + /H + and Cl – /HCO 3 – exchange. However, an important factor may be a lingering rise in Ca i 2+ in cells whose pH i is reduced over a longer period, predisposing to Ca 2+ -related damage. The molecular mechanisms underlying delayed acidosis-related damage probably comprise Fe 2+ and NO · -related production of free radicals that have the microvessels as their main target.

A concept of space for building classification, product modelling, and design

Information about a buildings spaces is of interest in every stage of the construction and facility management processes. An organisation1 or enterprise is located in and uses the buildings spaces, and many of the buildings spatial properties are determined on the basis of the user organisations requirements. The definition of the concept “space” as applied in information systems for building classification and building product modelling today is unclear. A fundamental problem is to reconcile a material and construction method viewpoint with a space-centred viewpoint. In order to enable communication among actors and computer systems in the construction process, the concepts used in model development and the corresponding terms have to be formally defined and standardised. In this article, we analyse the concept of space and suggest a comprehensive definition for the construction context. The identification of a space in a building is based on a spatial view. We introduce the concept of aspectual unit and show how this concept can be used to integrate different aspect views in a conceptual schema. Additionally, we define the user organisation as a thing, which is separate from the building and has spatial properties of its own, so-called “activity spaces”. Finally, we show how space may be represented in a comprehensive conceptual schema.

Influence of hyperglycemia and of hypercapnia on cellular calcium transients during reversible brain ischemia

The object of the study was to find out how preischemic hyperglycemia (in normocapnic animals) or excessive hypercapnia (in normoglycemic animals) affect the calcium transient during ischemia, as this can be assessed by measurements of the extracellular calcium concentration ([Ca2+]e). To that extent, normocapnic-normoglycemic control animals were compared with animals with induced hyperglycemia or hypercapnia, all being subjected to 10 min of forebrain ischemia, the [Ca2+]e and d.c. potential being measured with ion-sensitive glass microelectrodes. Hyperglycemia and hypercapnia delayed the loss of ion homeostasis following induction of ischemia. Furthermore, both hyperglycemia and hypercapnia reduced the delay of Ca2+ extrusion upon recirculation. As a result, both hyperglycemia and hypercapnia significantly reduced the ischemic calcium transient, as this was assessed by calculating the duration of maximal calcium load of cells. The results make it less likely that aggravation of brain damage by hyperglycemia or excessive hypercapnia is related to a further derangement of cell calcium homeostasis.

Perturbation of cellular energy state in complete ischemia: Relationship to dissipative ion fluxes

SummaryLoss of cellular ion homeostasis during anoxia, with rapid downhill fluxes of K+, Ca2+, Na+ and Cl-, is preceded by a slow rise in extracellular K+ concentration (Ke+), probably reflecting early activation of a K+ conductance. It has been proposed that this conductance is activated by either a rise in intracellular calcium concentration (Cai2+), or by a fall in ATP concentration. In a previous study from this laboratory (Folbergrová et al. 1990) we explored whether the early activation of a K+ conductance could be triggered by a rise in Cai2+. To that end, labile metabolites and phosphorylase a, a calcium sensitive enzyme, were measured after 15, 30, 60 and 120 s of complete ischemia (“anoxia”). In the present study, we investigated whether brief anoxia is accompanied by changes in ATP/ADP ratio, or in the phosphate potential, which could cause activation of a K+ conductance. To provide information on this issue, we added a group with 45 s of anoxia to the previously reported groups, and derived changes in intracellular pH (pHi). This allowed calculations of the free concentrations of ADP (ADPf) and AMP (AMPf) from the creatine kinase and adenylate kinase equilibria, and hence the derivation of ATP/ADPf ratios. In performing these calculations we initially assumed that the free intracellular Mg2+ concentration remained unchanged at 1 mM. However we also explored how a change in Mgi2+of the type described by Brooks and Bachelard (1989) influenced the calculation. The results showed that ADPf must have risen to 150–200% of control within 15 s, and to 330–350% of control within 45 s of anoxia. The concentration of AMPf should have increased 2–4 fold in 15 s and 10–20 fold in 45 s. Thus although tissue ATP concentration usually remains >90% of control within the first 30s of anoxia, and >80% of control within the first 45 s, the ATP/ADPf ratios change markedly at a time when alterations in ion homeostasis are dominated by a moderate rise in Ke+, and long before massive ion fluxes occur and the cells depolarise (after about 60–70 s). Such early changes in ATP/ADPf ratio, or in phosphate potential, could well influence reactions which are coupled to ATP hydrolysis, and perhaps lead to activation of ATP-dependent K+ conductances.

Coupling of Energy Failure and Dissipative K+ Flux during Ischemia: Role of Preischemic Plasma Glucose Concentration

The present experiments were undertaken to assess the influence of preischemic hypo- or hyperglycemia on the coupling among changes in extracellular K+ concentration (K+e) and in cellular energy state, as the latter is reflected in the tissue concentrations of phosphocreatine (PCr), Cr, ATP, ADP, and AMP, and in the calculated free ADP (ADPf) concentrations. The questions posed were whether the final release of K+ was delayed because the extra glucose accumulated by hyperglycemic animals produced enough ATP to continue supporting Na+–K+-driven ATPase activity, and whether the additional acidosis altered the ionic transients. As expected, preischemic hypoglycemia shortened and hyperglycemia prolonged the phase before K+e rapidly increased. This was reflected in corresponding changes in tissue ATP content. Thus, hypoglycemia shortened and hyperglycemia prolonged the time before the fall in ATP concentration accelerated. When tissue was frozen at the moment of depolarization, the tissue contents of ATP were similar in hypo-, normo-, and hyperglycemic groups, ∼ 30% of control. This suggests that hyperglycemia retards loss of ion homeostasis by leading to production of additional ATP. However, hyperglycemia did not reduce the rate at which the PCr concentration fell, and the ATP/ADPf ratio decreased. There were marked differences in the amount of lactate accumulated between the groups. Thus, massive depolarization in hypoglycemic groups occurred at a tissue lactate content of ∼4 mM kg−1. This corresponds to a decrease in intracellular pH (pHi) from ∼7.0 to ∼6.9. In the hyperglycemic groups, depolarization occurred at a lactate content of about 12 mm kg−1, corresponding to a pHi of ∼6.4. This fall in pHi, or the accompanying fall in extracellular pH (pHe), did not affect the maximal rate of efflux of K+. Measurements of ischemic depolarization at constant tissue temperature (37°C) suggest that the influence of the plasma glucose concentration on the terminal depolarization time is restricted. Thus, the time to depolarization varied between 30 s (hypoglycemia) and 90 s (moderate to severe hyperglycemia). Previous results obtained without temperature control may well have reflected a combination between hyperglycemia and a fall in tissue temperature.

Tissue Pco2 in Brain Ischemia Related to Lactate Content in Normo- and Hypercapnic Rats

The amount of lactate formed during ischemia determines the rise in tissue Pco2 (Ptco2). Conflicting results exist on the relationship between lactate and Ptco2. The objective of this study was to settle this issue. We varied the preischemic plasma glucose concentration of normo- and hypercapnic rats, assessed tissue lactate and total CO2 contents, and determined the Pco2/lactate relationship over the lactate range 2–40 mmol kg−1. The results showed that whatever the equilibration time, the Pco2/lactate relationship was linear. The results obtained could be reproduced by a theoretical buffer system that mimics the buffering behavior of intracellular fluid. Our results bear on the question of whether compartmentation of H+ occurs during ischemia, with glial cells becoming more acid than neurons. A discontinuous Pco2/lactate relationship, with a constant Pco2 above a certain lactate content, would support this contention. Since our results demonstrate a linear relationship between lactate and Pco2 over the lactate range 2–40 mmol kg−1, they considerably weaken any argument for gross compartmentation of H+.