Caffeine-Induced Ca2+ Transients and Exocytosis in Paramecium Cells. A Correlated Ca2+ Imaging and Quenched-Flow/Freeze-Fracture Analysis |
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Authors: | N Klauke H Plattner |
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Institution: | (1) Faculty of Biology, University of Konstanz, P.O. Box 5560, D-78434 Konstanz, Germany, DE |
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Abstract: | Caffeine causes a Ca2+]
i
increase in the cortex of Paramecium cells, followed by spillover with considerable attenuation, into central cell regions. From Ca2+]rest
i
∼50 to 80 nm, Ca2+]act
i
rises within ≤3 sec to 500 (trichocyst-free strain tl) or 220 nm (nondischarge strain nd9–28°C) in the cortex. Rapid confocal analysis of wildtype cells (7S) showed only a 2-fold cortical
increase within 2 sec, accompanied by trichocyst exocytosis and a central Ca2+ spread during the subsequent ≥2 sec. Chelation of Ca2+
o
considerably attenuated Ca2+]
i
increase. Therefore, caffeine may primarily mobilize cortical Ca2+ pools, superimposed by Ca2+ influx and spillover (particularly in tl cells with empty trichocyst docking sites). In nd cells, caffeine caused trichocyst
contents to decondense internally (Ca2+-dependent stretching, normally occurring only after membrane fusion). With 7S cells this usually occurred only to a small
extent, but with increasing frequency as Ca2+]
i
signals were reduced by Ca2+]
o
chelation. In this case, quenched-flow and ultrathin section or freeze-fracture analysis revealed dispersal of membrane components
(without fusion) subsequent to internal contents decondensation, opposite to normal membrane fusion when a full Ca2+]
i
signal was generated by caffeine stimulation (with Ca2+
i
and Ca2+
o
available). We conclude the following. (i) Caffeine can mobilize Ca2+ from cortical stores independent of the presence of Ca2+
o
. (ii) To yield adequate signals for normal exocytosis, Ca2+ release and Ca2+ influx both have to occur during caffeine stimulation. (iii) Insufficient Ca2+]
i
increase entails caffeine-mediated access of Ca2+ to the secretory contents, thus causing their decondensation before membrane fusion can occur. (iv) Trichocyst decondensation
in turn gives a signal for an unusual dissociation of docking/fusion components at the cell membrane. These observations imply
different threshold Ca2+]
i
-values for membrane fusion and contents discharge.
Received: 23 May 1997/Revised: 18 August 1997 |
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Keywords: | : Caffeine — Calcium — Exocytosis — Paramecium— Secretion |
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