Extracellular Potassium Levels Research Articles

Polydiacetylene Liposome Arrays for Selective Potassium Detection

Potassium is an important cation in biology, and quantitative detection of the extracellular potassium level is important. However, selective detection of extracellular physiological potassium is a challenging task due to the presence of sodium in a much higher concentration. In this contribution, we describe the development of practical polydiacetylene (PDA) liposome-based microarrays to selectively detect potassium even in the presence of sodium. We utilize the fact that the G-rich ssDNA can fold into a G-quadruplex via intramolecular hydrogen bonding by wrapping around a potassium ion exclusively. We rationally design the PDA liposome in such a way that the G-rich ssDNA probes are presented densely at the liposome surface and form bulky quadruplexes upon binding with K+. The resulting bulky quadruplexes are sterically hindered and repulse each other and impose mechanical stress on the PDA backbone, resulting in the conformational change of the ene-yne backbone of the PDA. As a result, polydiacetylene liposomes turn into the emissive red phase from the nonfluorescent blue phase.

The Back and Forth of Dendritic Plasticity

Synapses are located throughout the often-elaborate dendritic tree of central neurons. Hebbian models of plasticity require temporal association between synaptic input and neuronal output to produce long-term potentiation of excitatory transmission. Recent studies have highlighted how active dendritic spiking mechanisms control this association. Here, we review new work showing that associative synaptic plasticity can be generated without neuronal output and that the interplay between neuronal architecture and the active electrical properties of the dendritic tree regulates synaptic plasticity.

Changes in potassium concentration and haematocrit associated with cardiopulmonary bypass in paediatric cardiac surgery

A blood prime is frequently required for paediatric bypass surgery to maintain adequate haematocrit (Hct). However, stored blood can have high extracellular potassium levels and this study aims to investigate the effect of stored blood on the potassium concentration, both in the prime and subsequently in the patient after cardiopulmonary bypass (CPB) has been established. In neonatal surgery, the stored blood may be irradiated if there is a question of impaired immunity. Irradiation may cause a further increase in potassium levels. Blood-primed circuits prepared for 320 consecutive paediatric bypass cases were analysed for electrolyte levels, Hct and acid-base status before and immediately after establishment of CPB. Patients were divided into three groups according to body weight (<5kg, 5-10kg and > 10 kg) and both stored blood and irradiated blood primes were compared. The potassium concentration was above the physiological range in all bypass primes pre-CPB and was significantly higher when using irradiated blood (8.12 +/- 2.54 mmol/L versus 4.94 +/- 3.35 mmol/L, p < 0.0001). Despite this, on commencing CPB, the potassium level remained within the physiological range in the majority of patients (4.16 +/- 2.72mmol/L for stored blood prime and 4.55 +/- 1.01mmol/L for irradiated blood, p = 0.02). However, in smaller patients (<5 kg) who had irradiated blood prime potassium level > 7.0 mmol/L, there was resultant hyperkalaemia (5.60 +/-0.90 mmol/L) on commencing CPB, that returned to normal later. No adverse clinical events were associated with the hyperkalaemia. Hct was well maintained on CPB (22-25%) in all groups and was not related to patient weight. Blood primes result in high potassium concentrations in the prime fluid that is more severe if irradiated blood is used. The concentration is not sufficient to cause hyperkalaemia in the patients on commencing CPB except when irradiated blood prime is used in infants < 5 kg. Hct is well maintained in all patient groups with the use of blood prime.

Electrolytes Disturbances and Seizures

Electrolytes Disturbances and Seizures

A Modified Process for Preparation of Pathogen Inactivated RBC Using S-303 in Multiple Additive Solutions Maintains RBC Function and Reduces Immunologic Activity.

Background: Treatment of RBC with S-303 (acridine derivative) inactivates pathogens and leukocytes. The original process for S-303 treatment of RBC (oSRBC) for pathogen inactivation (PI) used 0.2mM S-303 and 2mM glutathione (GSH). Two randomized, controlled Phase III trials were voluntarily discontinued when antibodies, specific to residual acridine on oSRBC, were detected in 2 patients. Following this observation, a modified PI process using 0.2mM S-303 and 20mM GSH (mSRBC) was developed to reduce S-303 binding to the RBC surface which inactivates a broad spectrum of pathogens and maintains good RBC function.Aims: This study was designed to demonstrate the reduction in binding of S-303 to the RBC surface and preservation of in vitro RBC function with MSRBC in preparation for Phase I clinical trials.Methods: Leukoreduced RBC were prepared in Erythrosol, AS-3, AS-5, SAG-M or PAGGS-M additive solution. mSRBC were treated with 20mM GSH, pH 9.0, plus 0.2mM S-303 and incubated for 20 hours at RT. Post-treatment binding of S-303 to the RBC surface was assessed by gel agglutination test using a mouse monoclonal antibody specific to acridine. In vitro function was assayed pre- and 20 hours post-treatment, and weekly for 35 days (d) of storage. Parameters assessed for in vitro RBC function include intracellular ATP, 2,3-DPG, hemolysis, and extracellular potassium, glucose and lactate.Results: No agglutination was detected by gel column for mSRBC compared to 4+ readings for oSRBC indicating a reduction in S-303 bound to the RBC surface. While S-303 treatment results in higher initial ATP levels compared to pre-treatment, the mSRBC ATP level is maintained throughout 35d of storage. At 35d, oSRBC displayed glucose and lactate concentrations comparable to control RBC. In contrast, mSRBC had lower glucose and higher lactate concentrations than control RBC, consistent with observed higher ATP levels from increased glucose metabolism. After 35d of storage, hemolysis for oSRBC, Erythrosol mSRBC, AS-5 mSRBC, SAG-M mSRBC, and PAGGS-M mSRBC was comparable to controls whereas AS-3 mSRBC hemolysis was similar with the exception of 1 donor in AS-3 that showed a higher hemolysis level. Extracellular potassium levels were unaffected by oSRBC and mSRBC treatment.Conclusion: A modified PI process has been developed for RBC that significantly reduces S-303 bound to the RBC surface compared to the original process while retaining the critical RBC functions. The reduction in bound S-303 offers the potential for PI treatment for RBC without immunologic reactivity.ATP (μmol/g Hb)% HemolysisGATMean (SD)Mean (SD)oSRBCt=03.9 (0.2)0.1 (0.0)t=20h4.8 (0.3)0.2 (0.0)4+35 day2.6 (0.4)0.4 (0.0)mSRBC Erythrosolt=03.4 (0.2)0.1 (0.0)t=20h5.7 (0.1)0.1 (0.0)035 day3.8 (0.2)0.1 (0.0)mSRBC AS-3t=03.4 (0.5)0.2 (0.1)t=20h6.2 (0.4)0.3 (0.1)035 day3.7 (0.4)0.7 (0.8)mSRBC AS-5t=03.3 (0.2)0.1 (0.1)t=20h5.4 (0.7)0.3 (0.4)035 day3.3 (0.4)0.5 (0.3)mSRBC SAG-Mt=03.10.0t=20h5.00.0035 dayin progressin progressmSRBC PAGGS-Mt=03.10.0t=20h5.10.0035 dayin progressin progress

A role for glutamate and glia in the fast network oscillations preceding spreading depression

The mechanism of the propagation of spreading depression is unclear. Classical theories proposed a self-maintained cycle fed by elevated potassium and/or glutamate in the extracellular space. Earlier we found in vivo a characteristic oscillatory field activity that is synchronous in a strip of tissue ahead of the oncoming wave of neuron depolarization and that occurs before the extracellular potassium level begins to rise [Herreras O, Largo C, Ibarz JM, Somjen GG, Marrín del Río R (1994) Role of neuronal synchronizing mechanisms in the propagation of spreading depression in the in vivo hippocampus. J Neurosci 14:7087–7098]. We investigated here the possible participation of glutamate and the role of glia in the prodromal field oscillations using extra and intracellular recordings and pharmacological manipulations in rat hippocampal slices. As earlier shown in vivo, field oscillations propagated ahead of the negative potential shift covering distances of up to 1 mm. The oscillatory prodromals were initially subthreshold but then each wave became crowned by a population spike. The frequency of the oscillatory prodromals was variable among slices (80–115 Hz), but constant in individual slices. The blockade of ionotropic glutamate receptors decreased the frequency of prodromal oscillations, retarded spreading depression propagation, and shortened the duration of depolarization. Blocking the glutamate membrane transport increased the oscillatory frequency. The selective metabolic poisoning of astrocytes led to gradual disorganization of prodromal oscillations whose frequency first increased and then decreased. Also, the amplitude of the population spikes within the burst diminished as individual cells fired fewer action potentials, although still phase-locked with population spikes. The effects of glial metabolic impairment were observed within the period when neuron electrical properties were still normal, and were blocked by glutamate receptor antagonists. These data suggest that glutamate released from glial cells and possibly also from neurons has a role in the generation of oscillations and neuron firing synchronization that precede the spreading depression-related depolarization, but additional mechanisms are required to fully explain the onset and propagation of spreading depression.

Comparison of X‐ray vs. gamma irradiation of CPDA‐1 red cells

Transfusion-associated graft-versus-host disease (TA-GVHD) is a serious, potential complication of blood transfusion that is essentially prevented by blood product irradiation. Blood product irradiation is currently performed using gamma irradiation. X-ray irradiation is an alternative that has certain advantages: an X-ray irradiator is less expensive and does not have a radioactive source. However, the biochemical effects of X-ray irradiation on red blood cells (RBCs) have not been well characterized. The primary purpose of our study was to compare the effects of X-ray irradiation with gamma irradiation on RBC membrane permeability. A secondary purpose was to verify that X-ray irradiation has the same effect on lymphocytes as gamma irradiation. Ten RBC units were split into two portions; five units of each were then irradiated with either gamma or X-ray at a dose of 35 Gy or 25 Gy, respectively. Free plasma haemoglobin and extracellular potassium levels were measured in each unit over time. Another unit was divided into three parts for lymphocyte studies; one part was not irradiated as a control and the other two received 25 Gy of irradiation via X-ray or gamma irradiation, respectively. Minimally increased free plasma haemoglobin was found only in X-ray-irradiated units after 25 Gy compared to gamma-irradiated units. However, extracellular potassium levels were not significantly different in these groups. Extracellular potassium levels were slightly higher only in gamma irradiated units at 35 Gy. X-ray and gamma irradiation were equivalent in affecting lymphocyte function. Small differences in RBC membrane permeability are found between gamma-irradiated and X-ray-irradiated units. However, these differences are not likely to be clinically important.

Electrocardiographic manifestations: electrolyte abnormalities

Because myocyte depolarization and repolarization depend on intra- and extracellular shifts in ion gradients, abnormal serum electrolyte levels can have profound effects on cardiac conduction and the electrocardiogram (EKG). Changes in extracellular potassium, calcium, and magnesium levels can change myocyte membrane potential gradients and alter the cardiac action potential. These changes can result in incidental findings on the 12-lead EKG or precipitate potentially life-threatening dysrhythmias. We will review the major electrocardiographic findings associated with abnormalities of the major cationic contributors to cardiac conduction—potassium, calcium and magnesium.

The Influence of Extracorporeal Circulation on the Susceptibility of Erythrocytes to Oxidative Stress

Extracorporeal circulation (ECC), a necessary and integral part of cardiac surgery, can itself induce deleterious effects in patients. The pathogenesis of diffuse damage of several tissues is multifactorial. It is believed that circulation of blood extracorporeally through plastic tubes causes a whole body inflammatory response and a severe shear stress to blood cells. The aim of this study was to evaluate the level of oxidative stress and its deleterious effect on red blood cell (RBC) before (pre-ECC), immediately after (per-ECC) and 24 h after an ECC (24 h post-ECC). Several indicators of extracellular oxidative status were evaluated. The ascorbyl free radical (AFR) was directly measured in plasma using electron spin resonance (ESR) spectroscopy and expressed with respect to vitamin C levels in order to obtain a direct index of oxidative stress. Allophycocyanin assay was also used to investigate the plasma antioxidant status (PAS). Indirect parameters of antioxidant capacities of plasma such as vitamin E, thiol and uric acid levels were also quantified. RBC alterations were evaluated through potassium efflux and carbonyl levels after action of AAPH, a compound generating carbon centered free radicals. No changes in plasma uric acid and thiols levels were observed after ECC. However, vitamin E levels and PAS were decreased in per-ECC and 24 h post-ECC samples. Vitamin C levels were significantly lower in 24 h post-ECC and the AFR/ vitamin C ratio was increased. Differences in results had been noted when measurements took account of hemodilution. Increases of uric acid and thiols levels were observed after ECC. Vitamin E levels were not modified. However after hemodilution correction a significant decrease of vitamin C level was noted in 24 h post-ECC samples as compared to per-ECC sample. Whatever the way of measurement, vitamin C levels decreased suggesting the occurrence of ECC induced-oxidative stress. Concerning RBC, in the absence of AAPH, extracellular potassium remained unchanged between pre-, per- and 24 h post-ECC. AAPH induced a significant increase in extracellular potassium and carbonyls levels of RBC membranes, which was not modified by ECC. These results suggest the absence of alterations of RBC membrane during ECC despite the occurrence of disturbances in PAS. Such protection is of particular importance in a cell engaged in the transport of oxygen and suggests that RBC are equipped with mechanisms affording a protection against free radicals.

Astrocytes contribute to regulation of extracellular calcium and potassium in the rat cerebral cortex during spreading depression

This study used spreading depression (SD), which is characterized by redistribution of ions, to examine the role of astrocytes in the regulation of extracellular potassium ([K + ] o ) and calcium ([Ca 2+ ] o ) levels. Recurrent spreading depression episodes were induced by application of 3 M potassium chloride to the cortex of adult anesthetized rats while monitoring the extracellular direct current (DC) potential shifts and changes in [K + ] o or [Ca 2+ ] o 6–7 mm away. The reversible glial toxins, fluorocitrate (FC) and fluoroacetate (FA), were injected locally into the cortex at doses that are selective for reducing glial function. The peak changes and area under the curve for [K + ] o and [Ca 2+ ] o , recovery rate for [K + ] o , and interval between spreading depression episodes were measured before and at various times after administration of the toxins. Both fluorocitrate and fluroacetate slowed the recovery of the [K + ] o and altered the recovery of the [Ca 2+ ] o . Local injection of glutamate uptake inhibitors or barium had no effect on the peak changes in [K + ] o or the rate of recovery of the [K + ] o . The slowing of the recovery rate is consistent with the hypothesis that glial cells play a role in the return of [K + ] o to baseline after spreading depression in the cortex in vivo. The change in movement of calcium after administration of FC suggests that astrocytes normally extrude calcium during spreading depression, resulting in rapid recovery of the levels of [Ca 2+ ] o with an overshoot. These findings demonstrate that astrocytes contribute to the regulation of both potassium and calcium during and after a stress to the ionic homeostatic mechanisms.

Astrocytes contribute to regulation of extracellular calcium and potassium in the rat cerebral cortex during spreading depression

This study used spreading depression (SD), which is characterized by redistribution of ions, to examine the role of astrocytes in the regulation of extracellular potassium ([K +] o) and calcium ([Ca 2+] o) levels. Recurrent spreading depression episodes were induced by application of 3 M potassium chloride to the cortex of adult anesthetized rats while monitoring the extracellular direct current (DC) potential shifts and changes in [K +] o or [Ca 2+] o 6–7 mm away. The reversible glial toxins, fluorocitrate (FC) and fluoroacetate (FA), were injected locally into the cortex at doses that are selective for reducing glial function. The peak changes and area under the curve for [K +] o and [Ca 2+] o, recovery rate for [K +] o, and interval between spreading depression episodes were measured before and at various times after administration of the toxins. Both fluorocitrate and fluroacetate slowed the recovery of the [K +] o and altered the recovery of the [Ca 2+] o. Local injection of glutamate uptake inhibitors or barium had no effect on the peak changes in [K +] o or the rate of recovery of the [K +] o. The slowing of the recovery rate is consistent with the hypothesis that glial cells play a role in the return of [K +] o to baseline after spreading depression in the cortex in vivo. The change in movement of calcium after administration of FC suggests that astrocytes normally extrude calcium during spreading depression, resulting in rapid recovery of the levels of [Ca 2+] o with an overshoot. These findings demonstrate that astrocytes contribute to the regulation of both potassium and calcium during and after a stress to the ionic homeostatic mechanisms.

A xanthine-based KMUP-1 with cyclic GMP enhancing and K(+) channels opening activities in rat aortic smooth muscle.

1. KMUP-1 (1, 3, 5 mg kg(-1), i.v.), a xanthine derivative, produced dose-dependent sustained hypotensive and short-acting bradycardiac effects in anaesthetized rats. This hypotensive effect was inhibited by pretreatment with glibenclamide (5 mg kg(-1), i.v.). 2. In endothelium-intact or denuded aortic rings preconstricted with phenylephrine, KMUP-1 caused a concentration-dependent relaxation. This relaxation was reduced by endothelium removal, the presence of NOS inhibitor L-NAME (100 microM) and sGC inhibitors methylene blue (10 microM) and ODQ (1 microM). 3. The vasorelaxant effects of KMUP-1 was attenuated by pretreatment with various K(+) channel blockers TEA (10 mM), glibenclamide (1 microM), 4-AP (100 microM), apamin (1 microM) and charybdotoxin (ChTX, 0.1 microM). 4. Increased extracellular potassium levels (30 - 80 mM) caused a concentration-related reduction of KMUP-1-induced vasorelaxations. Preincubation with KMUP-1 (1, 10, 100 nM) increased the ACh-induced maximal vasorelaxations mediated by endogenous NO release, and enhanced the potency of exogenous NO-donor SNP. 5. The vasorelaxant responses of KMUP-1 (0.01, 0.05, 0.1 microM) together with a PDE inhibitor IBMX (0.5 microM) had an additive action. Additionally, KMUP-1 (100 microM) affected cyclic GMP metabolism since it inhibited the activity of PDE in human platelets. 6. KMUP-1 induced a dose-related increase in intracellular cyclic GMP levels in rat A10 vascular smooth muscle (VSM) cells, but not cyclic AMP. The increase in cyclic GMP content of KMUP-1 (0.1 - 100 microM) was almost completely abolished in the presence of methylene blue (10 microM), ODQ (10 microM), and L-NAME (100 microM). 7. In conclusion, these results indicate that KMUP-1 possesses the following merits: (1) stimulation of NO/sGC/cyclic GMP pathway and subsequent elevation of cyclic GMP, (2) K(+) channels opening, and (3) inhibition of PDE or cyclic GMP breakdown. Increased cyclic GMP display a prominent role in KMUP-1-induced VSM relaxations.

Induction of neuronal apoptosis by thiol oxidation: putative role of intracellular zinc release.

The membrane-permeant oxidizing agent 2,2'-dithiodipyridine (DTDP) can induce Zn(2+) release from metalloproteins in cell-free systems. Here, we report that brief exposure to DTDP triggers apoptotic cell death in cultured neurons, detected by the presence of both DNA laddering and asymmetric chromatin formation. Neuronal death was blocked by increased extracellular potassium levels, by tetraethylammonium, and by the broad-spectrum cysteine protease inhibitor butoxy-carbonyl-aspartate-fluoromethylketone. N,N,N', N'-Tetrakis-(2-pyridylmethyl)ethylenediamine (TPEN) and other cell-permeant metal chelators also effectively blocked DTDP-induced toxicity in neurons. Cell death, however, was not abolished by the NMDA receptor blocker MK-801, by the intracellular calcium release antagonist dantrolene, or by high concentrations of ryanodine. DTDP generated increases in fluorescence signals in cultured neurons loaded with the zinc-selective dye Newport Green. The fluorescence signals following DTDP treatment also increased in fura-2- and magfura-2-loaded neurons. These responses were completely reversed by TPEN, consistent with a DTDP-mediated increase in intracellular free Zn(2+) concentrations. Our studies suggest that under conditions of oxidative stress, Zn(2+) released from intracellular stores may contribute to the initiation of neuronal apoptosis.

Chronic hyperglycaemia increases neuronal sensitivity to high potassium in hippocampal slices from streptozotocin-treated diabetic rats

The chronic hyperglycaemia associated with diabetes mellitus increases susceptibility to epileptiform-like activity in the central nervous system. Changes in extracellular potassium levels directly influence neuronal excitability and significantly one of the major regulators of extracellular potassium, the Na–K ATPase, is known to be down-regulated by chronic hyperglycaemia. The sensitivity of hippocampal slices, from rats made diabetic for 2–3 weeks with streptozotocin, to increases in the extracellular potassium concentration ([K +] o) was, therefore, investigated. Raising [K +] o to 10 mM increased the number of orthodromically-evoked population spikes (PSs) in area CA1. This recruitment was significantly greater in hippocampal slices from diabetic rats, which also exhibited significantly more spontaneous activity. These findings confirm a diabetes-dependent increase in sensitivity of central neurones to changes in [K +] o and may help to explain the increased susceptibility to epileptiform activity in this disease state.

ECG changes during furosemide-induced hypokalemia in the rat

Electrolyte abnormalities have become an increasingly important cause of arrhythmias owing to the widespread use of high-potency diuretics. Hypokalemia is one of the common complications of diuretic use. Although some studies of hypokalemia induced by furosemide as well as of potassium-deficient diets in the rat have been reported, the electrocardiographic (ECG) changes during hypokalemia in the rat are poorly understood. This study was designed to examine such changes. For this purpose, hypokalemia was induced by furosemide administration, and the diagnostic criteria for ECG manifestations of hypokalemia were determined. During hypokalemia, conduction in most parts of the heart was suppressed to an extent depending on plasma potassium concentration. Prolongation of the QT interval was also observed, which agrees with findings in humans and dogs. Furthermore, prolonged durations of the P wave and QRS complex were observed during hypokalemia in the rat. The extent of alteration of the PR interval induced by hypokalemia was less significant than that of P wave and QRS complex durations. These results suggest that the excitabilities of the myocardium in the atria and ventricles may be affected by extracellular potassium level rather than by the atrioventricular conduction system in the rat. Wave amplitude, except that of the P wave, was decreased by severe hypokalemia. These changes were not dependent on the plasma potassium concentration. Typical T wave changes observed with hypokalemia in humans and dogs did not occur in the rat. The ECG manifestations of acute hypokalemia in the rat did not include the typical T wave changes seen in species with ST-segment type ECGs; however, other ECG parameter changes occurring with hypokalemia were qualitatively similar to those in other species. These results may be useful for testing the toxicity of potassium-depleting drugs in the rat.

Reinforcement, glucose metabolism and memory formation: A possible role for astrocytes

In the CNS, information processing, including memory formation, has been rightly viewed as primarily a neuronal function. In recent years, however, considerable evidence has been accumulated suggesting that astrocytes, a subclass of glia cells, may play a critical role in memory processing through their interactions with neurones in the regulation of the cellular microenvironment, in neurotransmitter and neuromodulator synthesis and release, and in energy metabolism. In day-old chicks trained on a single-trial discriminated passive avoidance task, astrocyte-neurone interactions in the regulation of extracellular potassium and calcium levels, in replenishment of neurotransmitter glutamate for glutamatergic and GABAergic neurones, and in glucose metabolism may determine whether a learning experience is consolidated into permanent memory. In particular, while the energy requirements for short-term memory stages are adequately met by glucose metabolism in neuronal and astrocytic citric acid cycles, this energy source appears to be insufficient for consolidation of permanent memory. Energy for the cellular processes underlying permanent memory appears to depend on the exclusively astrocytic metabolism of glycogen, stored in astrocytes but not in neurones. Furthermore, glycogenosis appears to be activated by the adequate release of noradrenaline, contingent on an appropriate level of reinforcement for the learning task.

Can the Indo-1 fluorescence approach measure brain intracellular calcium in vivo? A multiparametric study of cerebrocortical anoxia and ischemia

Indo-1 fluorescence was used to monitor intracellular calcium levels in the cat brain in vivo, using the approach proposed by Uematsu et al. [Uematsu D., Greenberg J.H., Reivich M., Karp A. In vivo measurement of cytosolic free calcium during cerebral ischemia and reperfusion. Ann Neurol 1988; 24: 420–428]. In addition, extracellular calcium and potassium levels, NADH redox state, electrocorticogram (ECoG), DC potential and relative cerebral blood flow were monitored simultaneously. Changes in the Indo-1 fluorescence ratio F 400/F 506 were monitored during anoxia, reversible ischemia and irreversible ischemia. Although these perturbations resulted in the expected changes in extracellular calcium and potassium levels, NADH redox state, ECoG and other physiological parameters, they did not result in significant increases in the F 400/F 506 ratio. The apparent insensitivity of the in vivo Indo-1 approach is due to the difficulty in obtaining accurate fluorescence signals from Indo-1 in the brain. Two reasons for this difficulty appear to be problems in loading Indo-1 into the brain, and problems in correcting Indo-1 fluorescence signals for changes in NADH fluorescence and changes in absorption of intrinsic chromophores. Under the conditions of our in vivo cat experiments, Indo-1 fluorescence is not a viable approach for measuring changes in cerebral intracellular calcium levels.

Neurosteroids modulate the reaction of astroglia to high extracellular potassium levels.

The extracellular concentration of potassium ([K+]o) in brain tissue is modified by neuronal activity and is increased under several pathological conditions. The influence of neurosteroids on the astroglia response to high [K+]o was assessed on cultured slices from rat hippocampus. Exposure to [K+]o above physiological (3 mM) levels resulted in the progressive appearance of cell processes immunoreactive for glial fibrillary acidic protein (GFAP). The maximal effect was observed at 50 mM [K+]o, and further increases of [K+]o did not increase the extension of GFAP-immunoreactive processes. The effect was observed as early as 10 min after increasing [K+]o, was independent of new protein synthesis, and was reversible, reaching control conditions by 15 h after resetting [K+]o to physiological levels. Gonadal hormones and neurosteroids had prominent and variable effects on the stimulatory influence of high [K+]o on astroglia morphology. At physiological [K+]o, 17beta-estradiol and pregnenolone, as well as its sulfate derivative, increased the extension of GFAP-immunoreactive processes. However, at high [K+]o, testosterone, pregnenolone, and dehydroepiandrosterone and its sulfate derivative decreased the extension of GFAP-immunoreactive processes. Effects of gonadal hormones and neurosteroids were blocked by the protein synthesis inhibitor cycloheximide. These results suggest that non-genomic effects of high [K+]o on glial cells interact with genomic effects of steroids to modulate astroglia morphology.

Neurosteroids modulate the reaction of astroglia to high extracellular potassium levels

The extracellular concentration of potassium ([K+]0) in brain tissue is modified by neuronal activity and is increased under several pathological conditions. The influence of neurosteroids on the astroglia response to high [K+]0 was assessed on cultured slices from rat hippocampus. Exposure to [K+]0 above physiological (3 mM) levels resulted in the progressive appearance of cell processes immunoreactive for glial fibrillary acidic protein (GFAP). The maximal effect was observed at 50 mM [K+]0, and further increases of [K+]0 did not increase the extension of GFAP-immunoreactive processes. The effect was observed as early as 10 min after increasing [K+]0, was independent of new protein synthesis, and was reversible, reaching control conditions by 15 h after resetting [K+]0 to physiological levels. Gonadal hormones and neurosteroids had prominent and variable effects on the stimulatory influence of high [K+]0 on astroglia morphology. At physiological [K+]0, 17β-estradiol and pregnenolone, as well as its sulfate derivative, increased the extension of GFAP-immunoreactive processes. However, at high [K+]0, testosterone, pregnenolone, and dehydroepiandrosterone and its sulfate derivative decreased the extension of GFAP-immunoreactive processes. Effects of gonadal hormones and neurosteroids were blocked by the protein synthesis inhibitor cycloheximide. These results suggest that non-genomic effects of high [K+]0 on glial cells interact with genomic effects of steroids to modulate astroglia morphology. © 1996 Wiley-Liss, Inc.

Three fluorescent probes for the flow-cytometric assessment of membrane potential in Saccharomyces cerevisiae.

Three fluorescent probes, tetramethyl rhodamine ethyl ester (TMRE), 3,3'-dipropylthiacarbocyanine iodide (diS-C3(3)) and 3,3'-dipropyloxacarbocyanine iodide (diO-C3(3)), were tested for their suitability as fluorescent indicators of membrane potential in Saccharomyces cerevisiae in studies performed by flow cytometry. For all these dyes the intensity of fluorescence of stained cells increased with probe concentration in the range of 60-3000 nmol/L. The optimum staining period was 15-20 min for diS-C3(3). Depolarization of cells by increased extracellular potassium level and by valinomycin elicited with all probes a drop in fluorescence intensity. In some yeast batches this depolarization was accompanied by a separation of subpopulations with different fluorescence properties.