Barium Evokes Glutamate Release from Rat Brain Synaptosomes by Membrane Depolarization: Involvement of K⁺, Na⁺, and Ca²⁺ Channels

Abstract

During K+-induced depolarization of isolated rat brain nerve terminals (synaptosomes), 1 mM Ba2+ could substitute for 1 mM Ca2' in evoking the release of endogenous glutamate. In addition, Ba2+ was found to evoke glutamate release in the absence of K+-induced depolarization. Ba2+ (1-10 mM) depolarized synaptosomes, as measured by voltage-sensitive dye fluorescence and r3H]-tetraphenylphosphonium cation distribution. Ba2+ partially inhibited the increase in synaptosomal K+ efflux produced by depolarization, as reflected by the redistribution of radiolabeled %b+. The release evoked by Ba2+ was inhibited by tetrodotoxin (TTX Guan et al., 1988;Matthies et al., 1988;Quastel and Saint, 1988;Heldman et al., 1989; Taglialatela et al., 1989; Hamano et al., 1991). By comparing and contrasting the action of Ba2+ with the action of Ca2+ on neurotransmitter release, important insights into the physiological mechanisms by which Ca2+ both directly triggers and indirectly regulates exocytosis may be revealed. One difference in the action of these two cations has, in fact, been noted previously: Ba2+, unlike Ca2+, was found to increase the release of neurotransmitter under resting (basal) conditions. Although Ba2+ has been shown to enter nerve terminal preparations largely through voltage-sensitive calcium channels (VSCCs; Nachshen and Blaustein, 1982; Augustine and Eckert, 1984), the nature of the Ba2+-induced increase in basal neurotransmitter release has not been clearly delineated.Two possibilities have been put forth to account for the effect of Ba2+ on neurotransmitter release. First, it has been suggested that following entry, Ba2+ accumulates in the nerve terminal cytosol to a greater extent than does Ca2+ (e.g., Quastel and Saint, 1988), because Ba2+ is not sequestered by the intraterminal buffering systems as effectively as Ca" (RasgadoFlores et al., 1985). Thus, even if the entry of Ba2+ under nondepolarizing conditions were low, Ba2+ would amass in the nerve terminal and induce release. Second, it has been suggested that Ba2+ acts by depolarizing nerve terminals through blockade of K+ channels (see Hagiwara et al., 1974), which would, in turn, open VSCCs and allow Ba2+ to enter. These possibilities are not mutually exclusive. Finally, once Ba2+ enters, it remains to be demonstrated in an intact nerve terminal that Ba" acts directly on the release Abbrevialions used: 4-AP, 4-arninopyndine; DiBAC4(3), bis( I ,3-dibutylbarbituric acid)tnmethine oxonol; HBS, HEPES-buffered saline; SDS, sodium dodecyl sulfate; TEA+, tetraethylammonium ion; TPP', tetraphenylphosphonium ion; TTX, tetrodotoxin; VSCC, voltage-sensitive calcium channels; WW78 1, bis(3-methyl-1 -psulphophenyl-5-pyrezolone)pentamethine oxonol.

Ba2+ replaces Ca2+/calmodulin in the activation of protein phosphatases and in exocytosis of all major transmitters

“…Thirdly, which cellular actions of Ba 2÷, if not calmodulin activation and calmodulin-dependent protein phosphorylation, are responsible for its stimulatory effect on transmitter release. The present results confirm that Ba 2+ ions, like Ca 2÷ ions, trigger transmitter release, as reported earlier (for instance Heldman et al, 1989;Tagliatela et al, 1989;Weiss et al, 1990;McMahon and Nicholls, 1993;Sihra et al, 1993). Ba 2+ and Ca 2+ were found to trigger release of all transmitters with similar kinetics and to a similar extent and to produce non-additive responses and thus appeared indistinguishable.…”

“…Ba 2+ ions exploit their permeability in Ca 2÷ channels and probably other, non-specific pathways to enter the terminal (Sihra et al, 1993). Ba 2÷ does not occur in high concentrations in the brain, has no known function in signal transduction in vivo, does not bind to many soluble Ca2+-buffering components and is sequestered within and/or extruded from the nerve terminal to a very limited extent (Chao et al, 1984;Rasgado-Flores et al, 1987;Mironov and Juri, 1990).…”

“…Although this process is reasonably specific for Ca2+, several divalent cations with similar physico-chemical properties, such as Ba'+, Sr2+ and Pb2+, were found to evoke transmitter release as well (Zengel and Magleby, 1980;Augustine and Eckert, 1984;Heldman et al, 1989;Shao and Suszkiw, 1991;McMahon and Nicholls, 1993;Sihra et al, 1993). Among these, Ba2+ is the best documented and probably the most effective.…”

“…Ba 2+ may depolarize the terminal by its unbalanced influx and by its inhibitory effect on K + channels (see Augustine and Eckert, 1984;Mironov and Juri, 1990;McMahon and Nicholls, 1993;Sihra et al, 1993). Intracellularly, Ba 2+ may exchange with Ca 2+ in internal stores.…”

“…Thirdly, which cellular actions of Ba 2+, if not calmodulin activation and calmodulin-dependent protein phosphorylation, are responsible for its stimulatory effect on transmitter release? Assay systems have been established to discriminate between indirect effects of Ba 2 + on the membrane potential, on intracellular Ca 2+ stores and on non-vesicular transmitter pools by using tetrodotoxin to reduce the instability of the membrane potential evoked by Ba 2+ (see McMahon and Nicholls, 1993;Sihra et al, 1993), by investigating additivity between Ba2+-evoked release and K+/Ca2+-evoked release and by exploiting both intact and permeated nerve terminals. In order to investigate the discrepancy between Ba2+-evoked transmitter release and the role of calmodulin in the same process, we have characterized the effects of the calmodulin inhibitor trifluoperizine on Ba 2+-and Ca2+-evoked release.…”