Biomedical subjects

S Morales-Mulia

Publications and source records attributed to S Morales-Mulia.

5 recordsLinked to original sources

PubMed1999

Influence of CA2+ on K+ efflux during regulatory volume decrease in cultured astrocytes.

The calcium (Ca2+) dependence of potassium (K+) efflux activated by hyposmolarity in cultured cerebellar astrocytes was investigated, measuring in parallel experiments (86)Rb release and changes in cytosolic Ca2+ ([Ca2+]i). Hyposmotic (50%) medium increased [Ca2+]i from 117 to 386 nM, with contributions of extracellular Ca2+ and Ca2+ from the endoplasmic reticulum. Hyposmotic medium increased (86)Rb efflux rate from 0.015 min(-1) to a maximal of 0. 049 min(-1) and a net release of 30%. This osmosensitive efflux was inhibited by Ba(2+) (0.028 min(-1)), quinidine (0.024 min(-1)), and charybdotoxin (0.040 min(-1)), but was unaffected by TEA, 4-AP, or apamin. Removal of external Ca2+ from the hyposmotic medium increased (86)Rb efflux to a maximal rate constant of 0.056 min(-1) and a net release of 38% and caused a delay of inactivation. These changes were due to the overlaping of an efflux activated by Ca2+ removal in isosmotic medium. This isosmotic 86Rb efflux was unaffected by TEA or 4-AP, reduced by verapamil, and abolished by Ba2+, nitrendipine, and Mg2+. With the swelling-induced [Ca2+]i rise suppressed by ethyleneglycoltetraacetic acid-acetoxy-methyl ester (EGTA-AM), hyposmotic (86)Rb was 30% reduced. The Ca2+ entry blockers Cd2+, Ni2+, La3+, and Gd3+ did not affect (86)Rb efflux. A 40% decrease observed with verapamil and nitrendipine was found unrelated to Ca2+, because these agents did not affect the [Ca2+]i rise and the inhibition persisted in the absence of external Ca2+. The phospholipase C blocker U-73122 did not affect [Ca2+]i nor (86)Rb efflux. Blockers of Ca2+/calmodulin W7 and KN-93 decreased (86)Rb efflux to the same extent as EGTA-AM. Ionomycin markedly potentiated (86)Rb release in hyposmotic conditions only when [Ca2+]i was raised to about 1 microM, suggesting the implication of maxi-K+ channels at this [Ca2+]i threshold, which nonetheless, was not attained during hyposmotic swelling. It is concluded that (86)Rb efflux in cerebellar astrocytes is largely (70%) Ca2+-independent and the Ca2+-dependent fraction is sustained essentially by Ca2+ released from the endoplasmic reticulum and mediated by a mechanism involving Ca2+/calmodulin.

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PubMed1998

Ca2+ changes and 86Rb efflux activated by hyposmolarity in cerebellar granule neurons.

Hyposmotic swelling increased 86Rb release in cultured cerebellar granule neurons (1 day in vitro [DIV]) with a magnitude related to the change in osmolarity. 86Rb release was partially blocked by quinidine, Ba2+, and Cs+ but not by TEA, 4-AP, or Gd3+. 86Rb efflux decreased in Cl(-)-depleted cells or cells treated with DDF or DIDS, suggesting an interconnection between Cl- and K+ fluxes. Swelling induced a substantial increase in [Ca2+]i to which both external and internal sources contribute. However, 86Rb efflux was independent of [Ca2+]0, unaffected by depleting the endoplasmic reticulum (ER) by ionomycin or thapsigargin and insensitive to charybdotoxin, iberiotoxin, and apamin. Swelling-activated 86Rb efflux in differentiated granule neurons after 8 DIV, which express Ca2+-sensitive K+ channels, was not different from that in 1 DIV neurons, nor in time course, net release, Ca2+-dependence, or pharmacological sensitivity. We conclude that the swelling-activated K+ efflux in cerebellar granule neurons is not mediated by Ca2+-sensitive large conductance K+ channels (BK) as in many cell types but resembles that in lymphocytes where it is possibly carried by voltage-gated K+ channels.

Animals↗

PubMed1998

Effects of NaCl removal on osmolyte fluxes and regulatory volume decrease in cultured astrocytes.

Cultured cerebellar astrocytes exhibit regulatory volume decrease (RVD) after anisosmotic swelling by extrusion of potassium (K), chloride (Cl), and organic osmolytes. The volume decay curve after swelling in 50% hyposmotic solutions has a first-order rate constant k min(-1) of 0.18 and 70% cell volume recovery is attained after 15 min. Replacing NaCl in the hyposmotic medium by sugars and polyalcohols markedly accelerated RVD increasing k to 0.37-0.39 min(-1) and full recovery in 3-5 min. NaCl removal increased (80%) K efflux (measured as 86Rb), delayed the inactivation of Cl efflux (measured as 125I) and enhanced taurine and D-aspartate release by 25%. These effects appear due to NaCl removal rather than to specific actions of the organic molecules. Replacing Na by other cations or Cl by gluconate increased k to 0.22 min(-1) and 0.26 min(-1), respectively, and 86Rb efflux by 4-23% and 39, respectively. It is concluded that the electrochemical gradient after NaCl removal increasing the K driving force, delaying the equilibrium of Cl and increasing amino acid efflux, accounts for the observed RVD acceleration. Other consequences of NaCl removal possibly modulating the K efflux pathway include changes in the surface charge screening, decreased ion strength, a rise in [Ca]i and a decrease in intracellular Na.

Amino Acids↗

PubMed1998

Osmotic swelling-induced changes in cytosolic calcium do not affect regulatory volume decrease in rat cultured suspended cerebellar astrocytes.

Hyposmotic swelling-induced changes in intracellular Ca2+ concentration ([Ca2+]i) and their influence on regulatory volume decrease (RVD) were examined in rat cultured suspended cerebellar astrocytes. Hyposmotic media (50 or 30%) evoked an immediate rise in [Ca2+]i from 117 nM to a mean peak increase of 386 (50%) and 220 nM (30%), followed by a maintained plateau phase. Ca2+ influx through the plasmalemma as well as release from internal stores contributed to this osmosensitive [Ca2+]i elevation. Omission of external Ca2+ or addition of Cd2+, Mn2+, or Gd3+ did not reduce RVD, although it was decreased by La3+ (0.1-1 mM). Verapamil did not affect either the swelling-evoked [Ca2+]i or RVD. Maneuvers that deplete endoplasmic reticulum (ER) Ca2+ stores, such as treatment (in Ca2+-free medium) with 0.2 microM thapsigargin (Tg), 10 microM 2,5-di-tert-butylhydroquinone, 1 microM ionomycin, or 100 microM ATP abolished the increase in [Ca2+]i but did not affect RVD. However, prolonged exposure to 1 microM Tg blocked RVD regardless of ER Ca2+ content or cytosolic Ca2+ levels. Ryanodine (up to 100 microM) and caffeine (10 mM) did not modify [Ca2+]i or RVD. BAPTA-acetoxymethyl ester (20 microM) abolished [Ca2+]i elevation without affecting RVD, but at higher concentrations BAPTA prevented cell swelling and blocked RVD. We conclude that the osmosensitive [Ca2+]i rise occurs as a consequence of increased Ca2+ permeability of plasma and organelle membranes, but it appears not relevant as a transduction signal for RVD in rat cultured cerebellar astrocytes.

Animals↗

PubMed1997

Regulatory volume decrease and associated osmolyte fluxes in cerebellar granule neurons are calcium independent.

To investigate a possible role for Ca as a transduction signal for regulatory volume decrease (RVD), the effects of external Ca removal, Ca channel blockers (Cd, Co, La, Gd, verapamil, diltiazem, dihydropyridines) and inhibitors of endoplasmic reticulum Ca release (dantrolene, ryanodine, TMB-8) were examined on RVD and on the swelling-activated efflux of two main osmolytes: Cl (traced by 125I) and [3H]taurine. Omission of Ca plus EGTA did not affect RVD or osmolyte release but when BAPTA was the chelator, RVD decreased 20%, 125I fluxes were unaffected and taurine stimulated efflux decreased (20%) while the basal efflux slightly increased (<10%). Verapamil, diltiazem, Co, Cd, La and Gd did not affect RVD or osmolyte fluxes. Nimodipine and nitrendipine (25-50 microM) markedly decreased RVD and osmolyte fluxes (>90%) through a mechanism independent of extracellular Ca. Swelling elicited an increase in cytosolic Ca measured by fura-2, which was notably variable ranging 50-350 nM. However, RVD and osmolyte fluxes were not affected by the blockers of endogenous Ca release dantrolene, ryanodine and TMB-8 or by the permeable Ca chelator BAPTA-AM, even when the cytosolic Ca increase was abolished by the chelator. These results indicate that 1) RVD and osmolyte fluxes are independent of extracellular Ca 2) RVD, osmolyte release and cytosolic Ca raise are only coincident events. Consequently, Ca is unlikely to be a transducing signal for RVD in neurons.

Animals↗