Occurrence of Na⁺-Ca²⁺ exchange in the ciliate Euplotes crassus and its role in Ca²⁺ homeostasis

Ciliates possess diverse Ca²⁺ homeostasis systems, but little is known about the occurrence of a Na⁺-Ca²⁺ exchanger. We studied Na⁺-Ca²⁺ exchange in the ciliate Euplotes crassus by digital imaging. Cells were loaded with fura-2/AM or SBF1/AM for fluorescence measurements of cytosolic Ca²⁺ and Na⁺ respectively. Ouabain pre-treatment and Na⁺(o) substitution in fura-2/AM-loaded cells elicited a bepridil-sensitive [Ca²⁺](i) rise followed by partial recovery, indicating the occurrence of Na⁺-Ca²⁺ exchanger working in reverse mode. In experiments on prolonged effects, ouabain, Na⁺(o) substitution, and bepridil all caused Ca²⁺(o)-dependent [Ca²⁺](i) increase, showing a role for Na⁺-Ca²⁺ exchange in Ca²⁺ homeostasis. In addition, by comparing the effect of orthovanadate (affecting not only Ca²⁺ ATPase, but also Na⁺-K⁺ ATPase and, hence, Na⁺-Ca²⁺ exchange) to that of bepridil on [Ca²⁺](i), it was shown that Na⁺-Ca²⁺ exchange contributes to Ca²⁺ homeostasis. In electrophysiological experiments, no membrane potential variation was observed after bepridil treatment suggesting compensatory mechanisms for ion effects on cell membrane voltage, which also agrees with membrane potential stability after ouabain treatment. In conclusion, data indicate the presence of a Na⁺-Ca²⁺ exchanger in the plasma membrane of E. crassus, which is essential for Ca²⁺ homeostasis, but could also promote Ca²⁺ entry under specific conditions.