scholarly journals Extracellular pH Changes and Accompanying Cation Shifts During Ouabain-Induced Spreading Depression

2000 ◽  
Vol 83 (3) ◽  
pp. 1338-1345 ◽  
Author(s):  
G. Menna ◽  
C. K. Tong ◽  
M. Chesler
Keyword(s):  
Spreading Depression ◽  
Cyclopiazonic Acid ◽  
Stratum Radiatum ◽  
Ph Changes ◽  
Alkaline Shift ◽  
Gradual Onset ◽  
High K ◽  
Ion Shifts ◽  
Free Media ◽  
Peak Value

Interstitial ionic shifts that accompany ouabain-induced spreading depression (SD) were studied in rat hippocampal and cortical slices in the presence and absence of extracellular Ca2+. A double-barreled ion-selective microelectrode specific for H+, K+, Na+, or Ca2+ was placed in the CA1 stratum radiatum or midcortical layer. Superfusion of 100 μM ouabain caused a rapid, negative, interstitial voltage shift (2–10 mV) after 3–5 min. The negativity was accompanied by a rapid alkaline transient followed by prolonged acidosis. In media containing 3 mM Ca2+, the alkalosis induced by ouabain averaged 0.07 ± 0.01 unit pH. In media with no added Ca2+ and 2 mM EGTA, the alkaline shift was not significantly different (0.09 ± 0.02 unit pH). The alkaline transient was unaffected by inhibiting Na+-H+ exchange with ethylisopropylamiloride (EIPA) or by blocking endoplasmic reticulum Ca2+ uptake with thapsigargin or cyclopiazonic acid. Alkaline transients were also observed in Ca2+-free media when SD was induced by microinjecting high K+. The late acidification accompanying ouabain-induced SD was significantly reduced in Ca2+-free media and in solutions containing EIPA. The ouabain-induced SD was associated with a rapid but relatively modest increase in [K+]o. In the presence of 3 mM external Ca2+, the mean peak elevation of [K+]o was 12 ± 0.62 mM. In Ca2+-free media, the elevation of [K+]o had a more gradual onset and reached a significantly larger peak value, which averaged 22 ± 1.1 mM. The decrease in [Na+]o that accompanied ouabain-induced SD was somewhat greater. The [Na+]o decreased by averages of 40 ± 7 and 33 ± 3 mM in Ca2+ and Ca2+-free media, respectively. In media containing 1.2 mM Ca2+, ouabain-induced SD was associated with a substantial decrease in [Ca2+]o that averaged 0.73 ± 0.07 mM. These data demonstrate that in comparison with conventional SD, ouabain-induced SD exhibits ion shifts that are qualitatively similar but quantitatively diminished. The presence of external Ca2+ can modulate the phenomenon but is irrelevant to the generation of the SD and its accompanying alkaline pH transient. Significance of these results is discussed in reference to the propagation of SD and the generation of interstitial pH changes.

scholarly journals Modulation of Spreading Depression by Changes in Extracellular pH

2000 ◽  
Vol 84 (5) ◽  
pp. 2449-2457 ◽  
Author(s):  
C. K. Tong ◽  
M. Chesler
Keyword(s):  
Nmda Receptor ◽  
Initial Velocity ◽  
Hippocampal Slice ◽  
Spreading Depression ◽  
Extracellular Ph ◽  
Stratum Radiatum ◽  
Acidic Media ◽  
Extracellular Acidosis ◽  
Alkaline Shift ◽  
Ph Microelectrodes

Spreading depression (SD) and related phenomena have been implicated in hypoxic-ischemic injury. In such settings, SD occurs in the presence of marked extracellular acidosis. SD itself can also generate changes in extracellular pH (pHo), including a pronounced early alkaline shift. In a hippocampal slice model, we investigated the effect of interstitial acidosis on the generation and propagation of SD in the CA1 stratum radiatum. In addition, a carbonic anhydrase inhibitor (benzolamide) was used to decrease buffering of the alkaline shift to investigate its role in the modulation of SD. pHo was lowered by a decrease in saline HCO3 − (from 26 to 13 to 6.5 mM at 5% CO2), or by an increase in the CO2 content (from 5 to 15% in 26 mM HCO3 −). Recordings with pH microelectrodes revealed respective pHo values of 7.23 ± 0.13, 6.95 ± 0.10, 6.67 ± 0.09, and 6.97 ± 0.12. The overall effect of acidosis was an increase in the threshold for SD induction, a decrease in velocity, and a shortened SD duration. This inhibition was most pronounced at the lowest pHo(in 6.5 mM HCO3 −) where SD was often blocked. The effects of acidosis were reversible on return to control saline. Benzolamide (10 μM) caused an approximate doubling of the early alkaline shift to an amplitude of 0.3–0.4 U pH. The amplified alkalosis was associated with an increased duration and/or increased velocity of the wave. These effects were most pronounced in acidic media (13 mM HCO3 −/5% CO2) where benzolamide increased the SD duration by 55 ± 32%. The initial velocity (including time for induction) and propagation velocity (measured between distal electrodes) were enhanced by 35 ± 25 and 26 ± 16%, respectively. Measurements of [Ca2+]o demonstrated an increase in duration of the Ca2+ transient when the alkaline shift was amplified by benzolamide. The augmentation of SD caused by benzolamide was blocked in media containing the N-methyl-d-aspartate (NMDA) receptor antagonistdl−2-amino-5-phosphonovaleric acid. These data indicate that the induction and propagation of SD is inhibited by a fall in baseline pH characteristic of ischemic conditions and that the early alkaline shift can remove this inhibition by relieving the proton block on NMDA receptors. Under ischemic conditions, the intrinsic alkalosis may therefore enable SD and thereby contribute to NMDA receptor-mediated injury.

Endogenous pH Shifts Facilitate Spreading Depression by Effect on NMDA Receptors

1999 ◽  
Vol 81 (4) ◽  
pp. 1988-1991 ◽  
Author(s):  
C. K. Tong ◽  
M. Chesler
Keyword(s):  
Nmda Receptor ◽  
Carbonic Anhydrase ◽  
Nmda Receptors ◽  
Spreading Depression ◽  
Hippocampal Slices ◽  
Nmda Receptor Antagonist ◽  
Ringer Solution ◽  
Stratum Radiatum ◽  
Alkaline Shift ◽  
Potential Electrode

Endogenous pH shifts facilitate spreading depression by effect on NMDA receptors. Rapid extracellular alkalinizations accompany normal neuronal activity and have been implicated in the modulation of N-methyl-d-aspartate (NMDA) receptors. Particularly large alkaline transients also occur at the onset of spreading depression (SD). To test whether these endogenous pH shifts can modulate SD, the alkaline shift was amplified using benzolamide, a poorly permeant inhibitor of interstitial carbonic anhydrase. SD was evoked by microinjection of 1.2 M KCl into the CA1 stratum radiatum of rat hippocampal slices and recorded by a proximal double-barreled pH microelectrode and a distal potential electrode. In Ringer solution of pH 7.1 containing picrotoxin (but not at a bath pH of 7.4), addition of 10 μM benzolamide increased the SD alkaline shift from 0.20 ± 0.07 to 0.38 ± 0.17 unit pH (means ± SE). This was correlated with a significant shortening of the latency and an increase in the conduction velocity by 26 ± 16%. In the presence of the NMDA receptor antagonist dl−2-amino-5-phosphonovaleric acid (APV), benzolamide still amplified the alkaline transient, however, its effect on the SD latency and propagation velocity was abolished. The intrinsic modulation of SD by its alkaline transient may play an important role under focal ischemic conditions by removing the proton block of NMDA receptors where interstitial acidosis would otherwise limit NMDA receptor activity.

Interstitial volume changes during spreading depression (SD) and SD-like hypoxic depolarization in hippocampal tissue slices

1994 ◽  
Vol 71 (6) ◽  
pp. 2548-2551 ◽  
Author(s):  
J. Jing ◽  
P. G. Aitken ◽  
G. G. Somjen
Keyword(s):  
Spreading Depression ◽  
Volume Changes ◽  
Tissue Slices ◽  
Ion Concentrations ◽  
Ca1 Region ◽  
Interstitial Volume ◽  
High K ◽  
Concentration Changes ◽  
Short Time ◽  
Total Tissue

1. Relative interstitial volume (ISV) was estimated from the concentration changes of iontophoretically administered tetramethyl- and tetraethylammonium (TMA+ and TEA+). Spreading depression (SD) was provoked by high K+, and hypoxic SD-like depolarization (HSD) was induced by withdrawing oxygen. 2. Probe ion concentrations increased dramatically and about equally during SD and HSD, except that in a few hypoxic trials signals became transiently smaller than control. Interstitial volume appeared to decrease on the average by approximately 70%. 3. The ISV that remains patent in CA1 region at the height of SD is < 4% of total tissue volume. Probe ions may occasionally have passed through cell membranes for a short time during hypoxic SD.

EFFECT OF IONIC ENVIRONMENT ON THE RELEASE OF HUMAN PLACENTAL LACTOGEN IN VITRO

1976 ◽  
Vol 69 (3) ◽  
pp. 349-358 ◽  
Author(s):  
V. J. CHOY ◽  
W. B. WATKINS
Keyword(s):  
Placental Tissue ◽  
Placental Lactogen ◽  
Human Placental Lactogen ◽  
Short Term ◽  
Graded Response ◽  
High K ◽  
Human Placental Tissue ◽  
High Concentrations ◽  
Free Media

SUMMARY Short-term incubation of human placental tissue in Krebs–Ringer bicarbonate buffered media with various concentrations of K+ and Ca2+ showed a graded response in human placental lactogen (HPL) release at different Ca2+ concentrations, but no effect at increased K+ concentration. Media with high Ca2+ caused an inhibition of release, while Ca2+-free media caused a stimulation in HPL release. High concentrations of Mg2+ inhibited release minimally, while Ba2+ had no effect. There was no change in HPL release when Na+ concentration was increased. La3+-Locke's solution markedly inhibited release of HPL but the significance of this effect is unknown. These results suggest that Ca2+ is not required for HPL secretion from placental tissue. It seems that HPL secretion in vitro does not follow the usual pattern where a physiological stimulus or high K+ concentration causes inward movement of calcium which couples stimulation to secretion.

scholarly journals Extracellular pH Changes during Spreading Depression and Cerebral Ischemia: Mechanisms of Brain pH Regulation

1984 ◽  
Vol 4 (1) ◽  
pp. 17-27 ◽  
Author(s):  
W. A. C. Mutch ◽  
A. J. Hansen
Keyword(s):  
Cerebral Ischemia ◽  
Parietal Cortex ◽  
Spreading Depression ◽  
Rapid Change ◽  
Extracellular Ph ◽  
Disulfonic Acid ◽  
Mammalian Brain ◽  
Negative Deflection ◽  
Alkaline Shift ◽  
Complete Cerebral Ischemia

We have examined the extracellular pH (pHe) during spreading depression and complete cerebral ischemia in rat parietal cortex utilizing double-barrelled H+ liquid ion exchanger microelectrodes. The baseline pHe of the parietal cortex was 7.33 at a mean arterial Pco2 of 38 mm Hg. Following spreading depression and cerebral ischemia, highly reproducible triphasic changes in pHe occurred, which were intimately related to the negative deflection in tissue potential (Ve). The changes in pHe for spreading depression (n = 23) were a small initial acidic shift, beginning before the rapid change in Ve, followed by a rapid transient alkaline shift of 0.16 pH units, the onset of which coincided with the negative deflection in Ve. A prolonged acidic shift of 0.42 pH units then occurred. The maximal decrease in pHe was to 6.97 and the mean duration of the triphasic pHe change was 7.8 min. The lactate concentration in brain cortex increased from baseline 1.2 m M to 7.0 m M (n = 6) during the maximal acidic change in spreading depression. In addition, lactate levels correlated well with resolution of the pHe changes during spreading depression. The triphasic pHe changes following complete cerebral ischemia were an initial acidic shift of 0.43 pH units which developed over 2 min, then an alkaline shift of 0.10 pH units coincident with the negative deflection in Ve, and a final acidic shift of 0.26 pH units. The terminal pHe was 6.75. Superfusion of the cortex with inhibitors of carbonic anhydrase (acetazolamide), Na+/H+ counter transport (amiloride), and Cl−/HCO3− countertransport (4,4'-diisothiocyanostilbene-2,2'-disulfonic acid) altered the triphasic pHe changes in a similar fashion for both spreading depression and cerebral ischemia, providing insights into the pHe regulatory mechanisms in mammalian brain.

Na+ pump α2-subunit expression modulates Ca2+ signaling

2003 ◽  
Vol 284 (2) ◽  
pp. C475-C486 ◽  
Author(s):  
Vera A. Golovina ◽  
Hong Song ◽  
Paul F. James ◽  
Jerry B. Lingrel ◽  
Mordecai P. Blaustein
Keyword(s):  
Plasma Membrane ◽  
Selective Reduction ◽  
Cyclopiazonic Acid ◽  
Cell Types ◽  
Membrane Microdomains ◽  
Α Subunit ◽  
Published Evidence ◽  
Subunit Expression ◽  
Mouse Fetuses ◽  
Free Media

The role of the Na+ pump α2-subunit in Ca2+ signaling was examined in primary cultured astrocytes from wild-type (α2 +/+ = WT) mouse fetuses and those with a null mutation in one [α2 +/− = heterozygote (Het)] or both [α2 −/− = knockout (KO)] α2 genes. Na+ pump catalytic (α) subunit expression was measured by immunoblot; cytosol [Na+] ([Na+]cyt) and [Ca2+] ([Ca2+]cyt) were measured with sodium-binding benzofuran isophthalate and fura 2 by using digital imaging. Astrocytes express Na+ pumps with both α1- (≈80% of total α) and α2- (≈20% of total α) subunits. Het astrocytes express ≈50% of normal α2; those from KO express none. Expression of α1 is normal in both Het and KO cells. Resting [Na+]cyt = 6.5 mM in WT, 6.8 mM in Het ( P > 0.05 vs. WT), and 8.0 mM in KO cells ( P < 0.001); 500 nM ouabain (inhibits only α2) equalized [Na+]cyt at 8 mM in all three cell types. Resting [Ca2+]cyt = 132 nM in WT, 162 nM in Het, and 196 nM in KO cells (both P < 0.001 vs. WT). Cyclopiazonic acid (CPA), which inhibits endoplasmic reticulum (ER) Ca2+ pumps and unloads the ER, induces transient (in Ca2+-free media) or sustained (in Ca2+-replete media) elevation of [Ca2+]cyt. These Ca2+ responses to 10 μM CPA were augmented in Het as well as KO cells. When CPA was applied in Ca2+-free media, the reintroduction of Ca2+ induced significantly larger transient rises in [Ca2+]cyt (due to Ca2+ entry through store-operated channels) in Het and KO cells than in WT cells. These results correlate with published evidence that α2 Na+ pumps and Na+/Ca2+ exchangers are confined to plasma membrane microdomains that overlie the ER. The data suggest that selective reduction of α2 Na+ pump activity can elevate local [Na+] and, via Na+/Ca2+ exchange, [Ca2+] in the tiny volume of cytosol between the plasma membrane and ER. This, in turn, augments adjacent ER Ca2+ stores and thereby amplifies Ca2+ signaling without elevating bulk [Na+]cyt.

scholarly journals Volume-dependent taurine release from cultured astrocytes requires permissive [Ca2+]iand calmodulin

1999 ◽  
Vol 277 (4) ◽  
pp. C823-C832 ◽  
Author(s):  
Alexander A. Mongin ◽  
Zhaohui Cai ◽  
Harold K. Kimelberg
Keyword(s):  
Cell Swelling ◽  
Transport Systems ◽  
Channel Blockers ◽  
Organic Osmolytes ◽  
Taurine Release ◽  
Cultured Astrocytes ◽  
High K ◽  
Intracellular Ca ◽  
Free Media ◽  
Taurine Efflux

Cell swelling results in regulatory activation of multiple conductive anion pathways permeable toward a broad spectrum of intracellular organic osmolytes. Here, we explore the involvement of extracellular and intracellular Ca2+ in volume-dependent [3H]taurine efflux from primary cultured astrocytes and compare the Ca2+ sensitivity of this efflux in slow (high K+ medium induced) and fast (hyposmotic medium induced) cell swelling. Neither Ca2+-free medium nor Ca2+-channel blockers prevented the volume-dependent [3H]taurine release. In contrast, loading cells with the membrane-permeable Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid (BAPTA)-AM suppressed [3H]taurine efflux by 65–70% and 25–30% under high-K+ and hyposmotic conditions, respectively. Fura 2 measurements confirmed that BAPTA-AM, but not Ca2+-free media, significantly reduced resting intracellular Ca2+concentration ([Ca2+]i). The calmodulin antagonists trifluoperazine and fluphenazine reversibly and irreversibly, respectively, inhibited the high-K+-induced [3H]taurine release, consistent with their known actions on calmodulin. In hyposmotic conditions, the effects were less pronounced. These data suggest that volume-dependent taurine release requires minimal basal [Ca2+]iand involves calmodulin-dependent step(s). Quantitative differences in Ca2+/calmodulin sensitivity of high-K+-induced and hyposmotic medium-induced taurine efflux are due to both the effects of the inhibitors on high-K+-induced cell swelling and their effects on transport systems and/or signaling mechanisms determining taurine efflux.

Effects of the Gliotoxin Fluorocitrate on Spreading Depression and Glial Membrane Potential in Rat Brain In Situ

1997 ◽  
Vol 78 (1) ◽  
pp. 295-307 ◽  
Author(s):  
Carlota Largo ◽  
José M. Ibarz ◽  
Oscar Herreras
Keyword(s):  
Membrane Potential ◽  
Rat Brain ◽  
Spreading Depression ◽  
Stratum Radiatum ◽  
Extracellular Potential ◽  
Neuronal Function ◽  
Neuron Death ◽  
Stratum Pyramidale ◽  
Glial Membrane

Largo, Carlota, José M. Ibarz, and Oscar Herreras. Effects of the gliotoxin fluorocitrate on spreading depression and glial membrane potential in rat brain in situ. J. Neurophysiol. 78: 295–307, 1997. DC extracellular potential shifts (Δ V o) associated with spreading depression (SD) reflect massive cell depolarization, but their cellular generators remain obscure. We have recently reported that the glial specific metabolic poison fluorocitrate (FC) delivered by microdialysis in situ caused a rapid impairment of glial function followed some hours later by loss of neuronal electrogenic activity and neuron death. We have used the time windows for selective decay of cell types so created to study the relative participation of glia and neurons in SD, and we report a detailed analysis of the effects of FC on evoked SD waves and glial membrane potential ( V m). Extracellular potential ( V o), interstitial potassium concentration ([K+]o), evoked potentials, and transmembrane glial potentials were monitored in the CA1 area before, during, and after administration of FC with or without elevated K+ concentration in the dialysate. SD waves propagated faster and lasted longer during FC treatment. Δ V o in stratum pyramidale, which normally are much shorter and of smaller amplitude than those in stratum radiatum, expanded during FC treatment to match those in stratum radiatum. The coalescing SD waves that develop late during prolonged high-K+ dialysis and are typically limited to stratum radiatum, also expanded into stratum pyramidale under the influence of FC. SD provoked in neocortex normally does not spread to the CA1, but during FC treatment it readily reached CA1 via entorhinal cortex. Once neuronal function began to deteriorate, SD waves became smaller and slower, and eventually failed to enter the region around the FC source. Slow, moderately negative Δ V o that mirrored [K+]o increments could still be recorded well after neuronal function and SD-associated V o had disappeared. Glial cell V m gradually depolarized during FC administration, beginning much before depression of neuronal antidromic action potentials. Calculations based on the results predict a large decrease in glial potassium content during FC treatment. The results are compatible with neurons being the major generator of the Δ V o associated with SD. We conclude that energy shortage in glial cells makes brain tissue more susceptible to SD and therefore it may increase the risk of neuron damage.

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