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

Abstract

Exocytosis from nerve terminals is triggered by depolarization-evoked Ca²⁺ entry, which also activates calmodulin and stimulates protein phosphorylation. Ba²⁺ is believed to replace Ca²⁺ in triggering exocytosis without activation of calmodulin and can therefore be used to unravel aspects of presynaptic function. We have analysed the cellular actions of Ba²⁺ in relation to its effect on transmitter release from isolated nerve terminals. Barium evoked specific release of amino acid transmitters, catecholamines and neuropeptides (EC₅₀ 0.2–0.5 mM), similar to K⁺/Ca²⁺-evoked release both in extent and kinetics. Ba²⁺- and Ca²⁺-evoked release were not additive. In contrast to Ca²⁺, Ba²⁺ triggered release which was insensitive to trifluoperizine and hardly stimulated protein phosphorylation. These observations are in accordance with the ability of Ba²⁺ to replace Ca²⁺ in exocytosis without activating calmodulin. Nevertheless, calmodulin appears to be essential for regular (Ca²⁺-triggered) exocytosis, given its sensitivity to trifluoperizine. Both Ba²⁺- and Ca²⁺-evoked release were blocked by okadaic acid. Furthermore, anti-calcineurin antibodies decreased Ba²⁺-evoked release. In conclusion, Ba²⁺ replaces Ca²⁺/calmodulin in the release of the same transmitter pool. Calmodulin-dependent phosphorylation appears not to be essential for transmitter release. Instead, our data implicate both Ca²⁺-dependent and -independent dephosphorylation in the events prior to neurotransmitter exocytosis.