Direct measurement of the effect of potassium,calcium, veratridine,and and amphetamine on the rate of release of dopamine from superfused brain tissue

The rate of release of endogenous DA from rat brain striatal minces has been measured using a rapid superfusion apparatus. The apparatus provides immediate, continuous readout of easily oxidized substances in the perfusate using an amperometric detector. Subsequent analysis of the perfusate (which contains pargyline) by liquid chromatography shows that the major substance detected is DA. DA release is induced by a 30 s exposure to 60 mM K⁺ and is Ca²⁺-dependent. Similar results are obtained with veratridine (10^?4 M). The time resolution of the perfusion system permits discrimination of the decreased rate of release induced by veratridine (10^?4 M) and amphetamine (10^?5 M) as opposed to 60 mM K⁺. Repetitive stimulation of the striatal mince with 60 mM K⁺ results in a decreased amount and rate of DA release. Subsequent exposure of the striatal mince to exogenous DA results in a restoration of the K⁺-induced, Ca²⁺-dependent release, indicating uptake of DA is operant under these conditions.     

Stimulation-evoked release of purines from the rabbit retina

The evoked release of purines from rabbit retinae preloaded with [³H]adenosine was studied in vitro. Potassium (8.6–43.6 mM) and ouabain (1 or 10 μM) increased the release of radioactivity in a concentration-dependent manner. The K⁺-evoked release was significantly reduced when the superfusion was carried out at 2–4°C. The effect of K⁺ (8.6, 13.6 and 23.6 mM) and of ouabain (1 μM) were completely abolished when the retinae were superfused with a Ca²⁺-free medium containing 0.1 mM EGTA. Calcium removal only partially reduced the effect of higher K⁺ and ouabain concentrations (43.6 mM and 10 μM, respectively). Further, the effect of K⁺ was found to be independent of extracellular Ca²⁺ when retinae were pretreated with ouabain for 30 min. Stimulation of the retina with light flashes induced a small, persistent increase in the release of radioactivity observable for several minutes after the end of stimulation.The superfusate contained mainly hypoxanthine and inosine. There were no significant changes in the relative proportions of the different purine compounds released before or in response to either K⁺ (23.6 mM) or ouabain (10 μM) stimulation. Potassium stimulation significantly increased the release of adenosine, inosine and hypoxanthine. Addition of the adenosine deaminase inhibitor, erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), significantly increased the relative proportions of released endogenous adenosine and inosine.The results indicate that K⁺ stimulation induces the release of purines from the rabbit retina by a Ca²⁺- and energy-dependent process. Light flashes also induce a purine release. The results suggest an active role for adenosine in retinal neurotransmission.     

Chloride dependence of the K+-stimulated release of taurine from synaptosomes

Exposure of a crude synaptosomal fraction to K⁺ concentrations ranging from 25 to 100 mM evokes the release of [³H]taurine and [³H]GABA. These high concentrations of K⁺ induce, besides depolarization, a marked synaptosomal swelling, which is prevented by replacing chloride in the solutions with the largely impermeant anion gluconate. The depolarizing effect of K⁺ is unaffected by omission of chloride. The K⁺-evoked release of taurine seems related to K⁺-induced changes in synaptosomal volume rather than to a depolarizing effect, since it is totally calcium-independent but is abolished by reducing chloride and by making solutions hypertonic with mannitol. The release of [³H]GABA, in contrast is unaffected in chloride-free or hypertonic solutions.     

Comparison of the effects of NaCl and KCl at the roots on seedling growth,cell death and the size,frequency and secretion rate of salt glands in leaves of <Emphasis Type="Italic">Limonium sinense</Emphasis>

Twenty days’ exposure to 50 or 100 mM NaCl in the rooting medium substantially increased fresh and dry weights of seedling shoots of the recretohalophyte Limonium sinense while 200 or 300 mM were increasingly inhibitory. KCl treatment was only slightly stimulating (50 mM) or strongly inhibitory (100–300 mM). Lesser effects on leaf area were also seen. Diameter of foliar salt glands was significantly larger than that of controls in 100 and 200 mM NaCl with the effect being reversed at higher concentrations. Gland enlargement was also observed in the presence of 100 mM KCl, while larger concentrations reduced gland size. Generally, gland diameter was larger in the presence of NaCl than in KCl. NaCl and KCl also increased gland number per leaf and secretion rate per gland. At 100 and 200 mM NaCl or KCl, Na⁺ secretion per leaf from NaCl-treated plants exceeded K⁺ secretion rate from KCl-treated plants while at 200 mM, Na⁺ secretion per gland was significantly higher for Na⁺ than for K⁺. Evidence of cell death in leaves of salt-treated plants using Evans blue staining indicates that release of cell contents through loss of membrane integrity contributed to the secretion values. We conclude that the greater tolerance of L. sinenseto to NaCl compared to KCl is linked to the more effective secretion of Na⁺ than of K⁺ and, in turn, to a greater stimulation of salt gland formation and activity and larger gland diameter.     

Alteration of cellular ionic constituents by external ionic conditions,and its significance in the development ofDictyostelium discoideum

Effects of external ionic conditions ofD. discoideum cells were examined in relation to intracellular ionic concentrations, the activity of pyruvate kinase and the amount of ATP. Main components of metal cations in heat extracts of vegetative cells were K⁺, Na⁺, Mg²⁺ and Ca²⁺ whose concentrations in a cell were about 35.0, 3.6, 10.6 and 2.3 mM, respectively. External Na⁺ at the concentration more than 50 mM inhibited the formation of cell aggregates in the presence of 10^?4M Ca²⁺. Such an inhibitory effect of Na⁺ was completely nullified by the addition of more than 10 mM K⁺. External Na⁺ caused a rapid decrease in intracellular K⁺, but an increase in intracellular Na⁺. Furthermore, it was found that the cells containing a high concentration of Na⁺ can develop normally in the presence of exogenous 10 mM K⁺, where intracellular K⁺ was maintaned at about 30 mM, irrespective of a high concentration of intracellular Na⁺ (about 30 mM). These suggest that the Na⁺-inhibition of the development is caused by a decrease in intracellular K⁺, but not by an increase in intracellular Na⁺. Pyruvate kinase extracted from the organism required K⁺ for its activation. The vegetative cells incubated in 50 mM Na⁺ contained only about 10 mM K⁺ which is insufficient for the enzyme activation. However, the amount of ATP in the cells containing less K⁺ was similar to that in those with much K⁺. These results are discussed in relation to the activity of glycolysis.     

Role of calcium in the thyrotropin-releasing hormone-stimulated release of prolactin from pituitary cells in culture.

Thyrotropin-releasing hormone (TRH) or 50 mM K⁺ stimulated the acute release of prolactin from the GH₄C₁ strain of rat pituitary cells in culture. The enhanced release of prolactin was inhibited in a dose-related manner by the Ca⁺² antagonist Co⁺² (2.0 to 0.5 mM) as well as by the Ca⁺² chelator EGTA (1.0 mM). Co⁺² also reduced spontaneous basal prolactin release. There was partial reversal of the inhibitory effect of Co⁺² (2.0 mM) by Ca⁺² (2.0 mM) and complete reversal of the inhibitory effect of EGTA (1.0 mM) by Ca⁺² (2.0 mM). The enhanced release of prolactin stimulated by 50 mM K⁺ was maximal by 10–20 minutes in medium containing 0.67 to 0.74 mM Ca⁺². Na⁺ (50 mM) did not mimic the effect of high K⁺. We conclude that Ca⁺² is an essential cation in mediating the actions of high external K⁺ and TRH on the release of prolactin by GH₄C₁ cells.     

Release of endogenous dopamine, 3,4-dihydroxyphenylacetic acid,and amino acid transmitters from rat striatal slices

The release of endogenous dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) was measured in superfused striatal slices of the rat and the results compared with data obtained for the release of endogenous (a) DA and DOPAC in the cerebral cortex, nucleus accumbens and thalamus; (b) 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), GABA, and glutamate in the striatum; and (c) GABA, glutamate and 5-HT in the cerebral cortex. In superfused slices of all four CNS regions, there appeared to be a Ca²⁺-dependent, K⁺-stimulated release of endogenous DA. In addition, in slices of the striatum and nucleus accumbens there also appeared to be a Ca²⁺-dependent, 60 mM K⁺ stimulated release of endogenous DOPAC. In the striatum, 16 mM Mg²⁺ was as effective as 2.5 mM Ca²⁺ in promoting the 60 mM K⁺-stimulated release of DOPAC. In addition, 16 mM Mg²⁺ appeared to function as a weak Ca²⁺ agonist since it also promoted the release of DA to approximately 40% of the level attained with Ca²⁺ in the presence of 60 mM K⁺. On the other hand, in the striatum, 16 mM Mg²⁺ inhibited the Ca²⁺-dependent, 60 mM K⁺-stimulated release of GABA and glutamate. Similar Mg²⁺-inhibition was observed in the cerebral cortex not only for GABA and glutamate but also for DA and 5-HT. With the use of -methyl -tyrosine (tyrosine hydroxylase inhibitor), cocaine (uptake inhibitor) and pargyline (monoamine oxidase inhibitor), it was determined that (a) most of the released DA and DOPAC was synthesized in the slices during the superfusion; (b) DOPAC was not formed from DA which had been released and taken up; and (c) DA and DOPAC were released from DA nerve terminals. In addition, the data indicate a difference in the release process between the amino acids and the monoamines from striatal slices since Mg²⁺ inhibited the Ca²⁺-dependent, K⁺-stimulated release of GABA and glutamate and appeared to promote the release of DA and 5-HT.     

EFFECT OF VINBLASTINE AND COLCHICINE ON UPTAKE AND RELEASE OF PUTATIVE TRANSMITTERS BY SYNAPTOSOMES AND ON BRAIN ACTOMYOSIN-LIKE PROTEIN

Abstract— The effects of several inhibitors, including vinblastine and colchicine, on the accumulation of a number of putative transmitters by a rat brain synaptosomal preparation and their subsequent release by excess K⁺ was examined. In addition, the effect of the alkaloids on the ATPase activity of the actomyosin-like protein, neurostenin, isolated from the synaptosomal preparation, was studied. The uptakes of radioactive glutamate, GABA, dopamine and norepinephrine were energy-dependent, as evidenced by their susceptibility to 0.01 mM carbonyl cyanide m-chlorophenylhydrazone (Cl-CCP), 01 mM ouabain and temperature. The active accumulations of GABA, dopamine and norepinephrine were also greatly inhibited by 1 mM6-hydroxydopamine (6-OHDA), 01 mM mersalyl, 0.05–0.25mM vinblastine and 0.1–1.0 mM colchicine. Vinblastine was approximately 10-fold more potent (K₁, ?0.1 mM) than colchicine as an inhibitor. The release of actively accumulated dopamine or norepinephrine by excess K⁺ (increasing the [K⁺] from 5 to 30 mM) was inhibited somewhat when vinblastine was present during the entire incubation period. If the synaptosomes were preloaded with the radioactive compounds prior to addition of vinblastine, there was no discernible effect on the relative amount of material released by excess K⁺. However, the addition of inhibitor under the latter conditions caused a leakage of radioactivity into the medium even without excess K⁺ being present. Glutamate accumulation was somewhat different from that of GABA, dopamine or norepinephrine. Although it required energy for uptake, 6-OHDA, mersalyl, vinblastine or colchicine were not inhibitory. Studies of the oxidative metabolism of glutamate and GABA by this synaptosomal preparation indicated that the mechanisms of inhibition by vinblastine was not attributable to a metabolic effect. Both vinblastine and colchicine inhibited the Mg²⁺-stimulated, but not the Ca²⁺-activated ATPase of neurostenin. This effect was probably attributable to an interaction of the vinblastine with the neurin moiety of this actomyosin-like protein. We suggest that the inhibitory phenomena exhibited by vinblastine and colchicine in this synaptosomal preparation arose from the effect of these alkaloids on the neurin associated with the synaptic membrane.     

Cloning and functional expression in <Emphasis Type="Italic">Saccharomyces cereviae</Emphasis> of a K<Superscript>+</Superscript> transporter,AlHAK, from the graminaceous halophyte, <Emphasis Type="Italic">Aeluropus littoralis</Emphasis>

High-affinity K⁺ transporters play an important role in K⁺ absorption of plants. We isolated a HAK gene from Aeluropus littoralis, a graminaceous halophyte. The amino acid sequence of AlHAK showed high homology with HAK transporters obtained from Oryza sativa (82%) and Hordeum vulgare (82%). When expressed in Saccharomyces cereviae WΔ3, AlHAK performed high-affinity K⁺ uptake with a K_m value of 8 μM, and the growth of transformants was dramatically inhibited by 150 mM Rb⁺ and 150 mM Cs⁺ but less affected by 300 mM Na⁺. AlHAK may thus improve the capacity of plants to maintain a high cytosolic K⁺/Na⁺ ratio at high salinity.     

K+-evoked taurine efflux from cerebellar astrocytes: On the roles of Ca2+ and Na+

The ionic requirements for K⁺-evoked efflux of endogenous taurine from primary cerebellar astrocyte cultures were studied. The Ca²⁺ ionophore A23187 evoked taurine efflux in a dose-dependent fashion with a time-course identical to that of K⁺-induced efflux. The Ca²⁺-channel antagonist nifedipine had no effect upon efflux induced by 10 or 50 mM K⁺. In addition, verapamil did not antagonize 50 mM K⁺-evoked efflux except at high, non-pharmacological concentrations (>100 M), and preincubation with 2 M -conotoxin had no effect on 50 mM K⁺-evoked efflux. Similarly, preincubation with 1 mM ouabain had no effect on the amount of taurine released by K⁺ stimulation, but did accelerate the onset of efflux by 2–4 min. Although 2 M tetrodotoxin had no effect on K⁺-evoked release, replacing Na⁺ with choline abolished the taurine efflux seen in response to K⁺ stimulation. Together, these findings suggest that neuronal N- and L-type Ca²⁺- and voltage-dependent Na⁺-channels are not involved in the influx of Ca²⁺ which appears to be necessary for K⁺-evoked taurine efflux, and that in addition to Ca²⁺, extracellular Na⁺ is also required.     

Muscimol Potentiation of Acidic Amino Acid Release from Cerebellar Synaptosomes Is Chloride Dependent

In previous studies we have shown that the depolarization-induced release of preaccumulated acidic amino acids and newly synthesized glutamate from cerebellar synaptosomal preparations is potentiated by γ-aminobutyric acid (GABA) agonists through a GABAergic presynaptic mechanism. Here we report a systematic analysis of the ionic requirements of the potentiating effect of muscimol on the high K⁺-evoked release of d-[³H]aspartate. Our studies show that: Ca²⁺, Na⁺, and Mg²⁺ are not required for muscimol to exert its effect; a depolarizing concentration of K⁺ is a necessary, but not sufficient, condition to observe the presynaptic effect in question; and a minimal Cl^- concentration (50–70 mM) is also required. A possible model based on these findings is proposed.     

K+ Uptake by Fermenting Escherichia coli Cells: pH Dependent Mode of the TrkA System Operating

Escherichia coli accumulates K⁺ by means of multiple transportsystems, of which TrkA is the most prominent at neutral and alkalinepH while Kup is major at acidic pH. In the present study, K⁺ uptakewas observed with cells grown under fermentative conditions at an initialpH of 9.0 and 7.3 (the medium pH decreased to 8.4 and 6.8, respectively,during the mid-logarithmic growth phase), washed with distilled water andresuspended in a K⁺ containing medium at pH 7.5 in the presence ofglucose. The kinetics for this K⁺ uptake and the amount of K⁺accumulated by the wild type and mutants having a functional TrkA orKup could confirm that K⁺ uptake by E. coli grown either at pH 9.0or pH 7.3 occurs mainly through TrkA. The following results distinguishpH dependent mode of TrkA operating: (1) K⁺ uptake was inhibited byDCCD in cells grown either at pH 9.0 or pH 7.3, although the stoichiometryof K⁺ influx to DCCD-inhibited H⁺ efflux for bacteria grownat pH 9.0 varied with external K⁺ concentration, but remained constantfor cells grown at pH 7.3; (2) K⁺ uptake was observed with an atpDmutant grown at pH 9.0 but not at pH 7.3; (3) The DCCD-inhibited H⁺efflux was increased 8-fold less by 5 mM K⁺ added into a K⁺ freemedium for bacteria grown at pH 9.0 than that for cells grown at pH 7.3;(4) the DCCD-inhibited ATPase activity of membrane vesicles from bacteriagrown at pH 9.0 was reduced a little in the presence of 100 mM K⁺,but stimulated more than 2.4-fold at pH 7.3.     

Modulation of Aspartate Release by Ascorbic Acid and Endobain E,an Endogenous Na<Superscript>+</Superscript>, K<Superscript>+</Superscript>-ATPase Inhibitor

The isolation of a soluble brain fraction which behaves as an endogenous ouabain-like substance, termed endobain E, has been described. Endobain E contains two Na⁺, K⁺-ATPase inhibitors, one of them identical to ascorbic acid. Neurotransmitter release in the presence of endobain E and ascorbic acid was studied in non-depolarizing (0 mM KCl) and depolarizing (40 mM KCl) conditions. Synaptosomes were isolated from cerebral cortex of male Wistar rats by differential centrifugation and Percoll gradient. Synaptosomes were preincubated in HEPES-saline buffer with 1 mM d-[³H]aspartate (15 min at 37°C), centrifuged, washed, incubated in the presence of additions (60 s at 37°C) and spun down; radioactivity in the supernatants was quantified. In the presence of 0.5–5.0 mM ascorbic acid, d-[³H]aspartate release was roughly 135–215% or 110–150%, with or without 40 mM KCl, respectively. The endogenous Na⁺, K⁺-ATPase inhibitor endobain E dose-dependently increased neurotransmitter release, with values even higher in the presence of KCl, reaching 11-times control values. In the absence of KCl, addition of 0.5–10.0 mM commercial ouabain enhanced roughly 100% d-[³H]aspartate release; with 40 mM KCl a trend to increase was recorded with the lowest ouabain concentrations to achieve statistically significant difference vs. KCl above 4 mM ouabain. Experiments were performed in the presence of glutamate receptor antagonists. It was observed that MPEP (selective for mGluR5 subtype), failed to decrease endobain E response but reduced 50–60% ouabain effect; LY-367385 (selective for mGluR1 subtype) and dizocilpine (for ionotropic NMDA glutamate receptor) did not reduce endobain E or ouabain effects. These findings lead to suggest that endobain E effect on release is independent of metabotropic or ionotropic glutamate receptors, whereas that of ouabain involves mGluR5 but not mGluR1 receptor subtype. Assays performed at different temperatures indicated that in endobain E effect both exocytosis and transporter reversion are involved. It is concluded that endobain E and ascorbic acid, one of its components, due to their ability to inhibit Na⁺, K⁺-ATPase, may well modulate neurotransmitter release at synapses.     

The influence of potassium content on the kinetics of potassium influx into excised ryegrass and barley roots

The influx of K⁺ into excised roots of barley (Hordeum vulgare L.) and ryegrass (Lolium multiflorum L.) previously grown with or without K⁺ was measured in K⁺ solutions ranging in concentration from 0.01 to 50 mM. In both species the K⁺ influx was lower in the roots with high K⁺ content. The extent of reduction by high internal [K⁺] decreased with external concentration above 1 mM. These results support the contention that at high external concentrations passive diffusion makes significant contributions to observed fluxes.     

Effects of High-Potassium-Induced Depolarization on Amino Acid Chemistry of the Dorsal Cochlear Nucleus in Rat Brain Slices

High K⁺ was used to depolarize glia and neurons in order to study the effects on amino acid release from and concentrations within the dorsal cochlear nucleus (DCN) of brain slices. The release of glutamate, -aminobutyrate (GABA) and glycine increased significantly during exposure to 50 mM K⁺, while glutamine and serine release decreased significantly during and/or after exposure, respectively. After 10 min of exposure to 50 mM K⁺, glutamine concentrations increased in all three layers of DCN slices, to more than 5 times the values in unexposed slices. In the presence of a glutamate uptake blocker, L-trans-pyrrolidine-2,4-dicarboxylic acid (PDC), glutamine concentrations in all layers did not increase as much during 50 mM K⁺. Similar but smaller changes occurred for serine. Mean ATP concentrations were lower in 50 mM K⁺-exposed slices compared to control. The results suggest that depolarization, such as during increased neural activity, can greatly affect amino acid metabolism in the cochlear nucleus.     

Comparison between electrically evoked and potassium-induced 3H-noradrenaline release from rat neocortex slices: Role of calcium ions and transmitter pools

In this study ³H-noradrenaline (NA) release from rat neocortex slices evoked by electrical field-stimulation (1 Hz, 12 mA, 2 msec) was compared with that induced by K⁺-depolarization (13–30 mM K⁺) under similar experimental conditions, with a particular emphasis on the role of external Ca²⁺ and the releasable transmitter pool(s). Not only ³H-NA release evoked by electrical stimulation but also that induced by 13 mM K⁺ was almost completely blocked by 0.3 μM tetrodotoxin (TTX). Release induced by 20 mM K⁺ appeared to be less sensitive to TTX. Thus, under relatively mild stimulation conditions, the activation of sodium channels appears to be involved in ³H-NA release elicited by both stimuli.The electrically evoked ³H-NA release increased sigmoidally with the external Ca²⁺-concentration up to 1.2 mM. In contrast, ³H-NA release induced by 13–20 mM K⁺ reached a maximal value at 0.6–0.9 mM Ca²⁺ and gradually decreased at higher Ca²⁺-concentrations. The Ca²⁺-antagonist D-600 (1–30 μM) did not inhibit electrically evoked release, while K⁺-induced ³H-NA release was dose-dependently reduced. Upon repetitive K⁺-depolarization a strong depression of ³H-NA release could be demonstrated, while this phenomenon did not occur with repeated electrical stimulation. Moreover, a previous K⁺-induced (partial) depletion of ³H-NA stores did not affect the release evoked by electrical pulses and vice versa. Taken together these data are compatible with a much stronger activation of Ca²⁺-channels and a larger vesicle mobilizing capacity in case of electrical stimulation at physiological frequencies compared to sustained depolarization with moderate K⁺-concentrations.     

Adaptation of Chinese hamster ovary cells to high potassium ion-containing medium for enhancement of follicle-stimulating hormone production

In this study, a suspension culture of recombinant Chinese hamster ovary (CHO) cells producing follicle-stimulating hormone (FSH) was used to investigate the effects of potassium ion (K⁺) on cell growth and FSH production. Cell growth was significantly suppressed at a K⁺ concentration higher than 60 mM, but specific FSH productivity (q _FSH) was enhanced more than 2-fold compared to the value obtained at 4 mM K⁺. In an attempt to alleviate the cell growth suppression at a high K⁺ concentration, the cells were adapted at 60 mM K⁺ in a repeated batch mode. During adaptation, the growth rate increased from 0.010 to 0.020 h⁻¹, andq _FSH also gradually increased and reached 11.1 ng/(10⁶ cells h), which was even higher than that of the unadapted cells at 60 mM K⁺. The adapted cells showed a 2.6-fold increase in maximum FSH titer at 80 mM K⁺ compared to the unadapted cells at 4 mM K⁺. Taken together, these results demonstrate the potential of using culture media containing cells adapted to high K⁺ concentrations, for the enhancement of recombinant protein production.     

Effects of potassium on the anion and cation contents of primary cultures of mouse astrocytes and neurons

Inastrocytes, as [K⁺]_o was increased from 1.2 to 10 mM, [K⁺]_i and [Cl^–]_i were increased, whereas [Na⁺]_i was decreased. As [K⁺]_o was increased from 10 to 60 mM, intracellular concentration of these three ions showed no significant change. When [K⁺]_o was increased from 60 to 122 mM, an increase in [K⁺]_i and [Cl^–]_i and a decrease in [Na⁺]_i were observed.Inneurons, as [K⁺]_o was increased from 1.2 to 2.8 mM, [Na⁺]_i and [Cl^–]_i were decreased, whereas [K⁺]_i was increased. As [K⁺]_o was increased from 2.8 to 30 mM, [K⁺]_i, [Na⁺]_i and [Cl^–]_i showed no significant change. When [K⁺]_o was increased from 30 to 122 mM, [K⁺]_i and [Cl^–]_i were increased, whereas [Na⁺]_i was decreased. Inastrocytes, pH_i increased when [K⁺]_o was increased. Inneurons, there was a biphasic change in pH_i. In lower [K⁺]_o (1.2–2.8 mM) pH_i decreased as [K⁺]_o increased, whereas in higher [K⁺]_o (2.8–122 mM) pH_i was directly related to [K⁺]_o. In bothastrocytes andneurons, changes in [K⁺]_o did not affect the extracellular water content, whereas the intracellular water content increased as the [K⁺]_o increased. Transmembrane potential (E_m) as measured with Tl-204 was inversely related to [K⁺]_o between 1.2 and 90 mM, a ten-fold increase in [K⁺]_o depolarized the astrocytes by about 56 mV and the neurons about 52 mV. The E_m values measured with Tl-204 were close to the potassium equilibrium potential (E_k) except those in neurons at lower [K⁺]_o. However, they were not equal to the chloride equilibrium potential (E_Cl) at [K⁺]_o lower than 30 mM in both astrocytes and neurons. Results of this study demonstrate that alteration of [K⁺]_o produced different changes in [K⁺]_i, [Na⁺]_i, [Cl^–]_i, and pH_i in astrocytes and neurons. The data show that astrocytes can adapt to alterations in [K⁺]_o, in such a way to maintain a more suitable environment for neurons.     

Modulation of [<Superscript>3</Superscript>H]Dopamine Release by Glutathione in Mouse Striatal Slices

Glutathione (γ-glutamylcysteinylglycine, GSH and oxidized glutathione, GSSG), may function as a neuromodulator at the glutamate receptors and as a neurotransmitter at its own receptors. We studied now the effects of GSH, GSSG, glutathione derivatives and thiol redox agents on the spontaneous, K⁺- and glutamate-agonist-evoked releases of [³H]dopamine from mouse striatal slices. The release evoked by 25 mM K⁺ was inhibited by GSH, S-ethyl-, -propyl-, -butyl- and pentylglutathione and glutathione sulfonate. 5,5′-Dithio-bis-2-nitrobenzoate (DTNB) and l-cystine were also inhibitory, while dithiothreitol (DTT) and l-cysteine enhanced the K⁺-evoked release. Ten min preperfusion with 50 μM ZnCl₂ enhanced the basal unstimulated release but prevented the activation of K⁺-evoked release by DTT. Kainate and 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) evoked dopamine release but the other glutamate receptor agonists N-methyl-d-aspartate (NMDA), glycine (1 mM) and trans-1-aminocyclopentane-1,3-dicarboxylate (t-ACPD, 0.5 mM), and the modulators GSH, GSSG, glutathione sulfonate, S-alkyl-derivatives of glutathione, DTNB, cystine, cysteine and DTT (all 1 mM) were without effect. The release evoked by 1 mM glutamate was enhanced by 1 mM GSH, while GSSG, glutathionesulfonate and S-alkyl derivatives of glutathione were generally without effect or inhibitory. NMDA (1 mM) evoked release only in the presence of 1 mM GSH but not with GSSG, other peptides or thiol modulators. l-Cysteine (1 mM) enhanced the glutamate-evoked release similarly to GSH. The activation by 1 mM kainate was inhibited by S-ethyl-, -propyl-, and -butylglutathione and the activation by 0.5 mM AMPA was inhibited by S-ethylglutathione but enhanced by GSSG. Glutathione alone does not directly evoke dopamine release but may inhibit the depolarization-evoked release by preventing the toxic effects of high glutamate, and by modulating the cysteine–cystine redox state in Ca²⁺ channels. GSH also seems to enhance the glutamate-agonist-evoked release via both non-NMDA and NMDA receptors. In this action, the γ-glutamyl and cysteinyl moieties of glutathione are involved.     

Basal and potassium stimulated, calcium dependent, endorphins release from pituitary cells.

Mouse pituitary tumor cells grown in tissue culture release endorphins spontaneously to the culture medium. Depolarization of these cells by incubation with high K⁺ concentration (56 mM) increased 2–3 folds the release of endorphins. The K⁺ evoked release was Ca⁺⁺ dependent by that: $\text{a}$, removal of Ca⁺⁺ ions inhibited 90% of K⁺ stimulated release. $\text{b}$, ethyleneglycol-bis (β-aminoethyl ether) N,N′-tetraacetic acid (EGTA) inhibited release of endorphins in the presence of high K⁺ and Ca⁺⁺. It is suggested that dual regulatory system inhibit and/or stimulate $\text{in-vivo}$ release of endorphins from the pituitary glands.