Ca2+ oscillations mediated by exogenous GTP in Paramecium cells: assessment of possible Ca2+ sources

Abstract

We applied exogenous guanosine trisphosphate, GTP, to Paramecium tetraurelia cells injected with Fura Red for analysing changes of free intracellular Ca 2+ concentrations, [Ca 2+ ] i , during periodic back-/forward swimming thus induced. Strain ginA (non-responsive to GTP) shows no Ca 2+ signal upon GTP application. In strain nd6 (normal Ca 2+ signalling) an oscillating [Ca 2+ ] i response with a prominent first peak occurs upon GTP stimulation, but none after mock-stimulation or after 15 min adaptation to GTP. While this is in agreement with previous electrophysiological analyses, we now try to identify more clearly the source(s) of Ca 2+ . Stimulation of nd6 cells, after depletion of Ca 2+ from their cortical stores (alveolar sacs), shows the same Ca 2+ oscillation pattern but with reduced amplitudes, and a normal behavioural response is observed. Stimulation with GTP, supplemented with the Ca 2+ chelator BAPTA, results in loss of the first prominent Ca 2+ peak, in reduction of the following Ca 2+ amplitudes, and in the absence of any behavioural response. Both these observations strongly suggest that for the initiation of GTP-mediated back-/forward swimming Ca 2+ from the extracellular medium is needed. For the maintenance of the Ca 2+ oscillations a considerable fraction must come from internal stores, probably other than alveolar sacs, rather likely from the endoplasmic reticulum.

Signalling in ciliates: long- and short-range signals and molecular determinants for cellular dynamics

“…When Paramecium tetraurelia cells are exposed to extracellular GTP, ≥10 µM, this causes periodic forward and backward swimming (different from ciliary reversal) in ∼8 s intervals; this is accompanied by oscillating membrane depolarisations (Clark, Hennessey & Nelson, ) and by oscillating intracellular Ca 2+ signals (Sehring & Plattner, ) that are consistent with oscillating Ca 2+ currents (Mimikakis, Nelson & Preston, ). This effect is specific to GTP and cannot be induced by any other nucleotide triphosphate in any comparable concentration.…”

“…This response to nucleotide triphosphates is unexpected insofar as there are no purinergic receptors known from ciliates (Burnstock & Verkhratsky, ; Coddou et al, ; Verkhratsky & Burnstock, ), in contrast to Dictyostelium discoideum (Fountain et al, ). In Paramecium tetraurelia GTP‐induced signalling involves release of Ca 2+ from unidentified internal stores (Wassenberg, Clark & Nelson, ), but a stronger first Ca 2+ peak depends on [Ca 2+ ] o (Sehring & Plattner, ). The biological significance of these Ca 2+ oscillations and of the repetitive back/forward movement is not quite clear, but it was suggested that the GTP‐induced behaviour counteracts dispersal of the cells (Sehring & Plattner, ).…”

“…In Paramecium tetraurelia GTP‐induced signalling involves release of Ca 2+ from unidentified internal stores (Wassenberg, Clark & Nelson, ), but a stronger first Ca 2+ peak depends on [Ca 2+ ] o (Sehring & Plattner, ). The biological significance of these Ca 2+ oscillations and of the repetitive back/forward movement is not quite clear, but it was suggested that the GTP‐induced behaviour counteracts dispersal of the cells (Sehring & Plattner, ).…”

Calcium Regulation in the Protozoan Model, Paramecium tetraurelia

“…). This is paralleled by periodic changes in [Ca 2+ ] i signals (Sehring and Plattner ). Based on inhibitor studies, the source of Ca 2+ underlying this signal was attributed to alveolar sacs (Wassenberg et al.…”

“…Considering that the pharmacology used is aberrant in Paramecium (Plattner et al. ) one may envisage some other stores which are important particularly for the first peak seen with a fluorochrome after GTP stimulation (Sehring and Plattner ). Which one of the many organelles endowed with CRCs (see below) this would be has not been determined as yet.…”