Imaging of Ca2+ transients induced in Paramecium cells by a polyamine secretagogue.
In Paramecium tetraurelia cells analysis of transient changes in Ca2+ concentration, [Ca2+]i, during aminoethyldextran (AED) stimulated synchronous (<1 second) trichocyst exocytosis has been hampered by various technical problems which we now have overcome. While Fura Red was found appropriate for quantitative double wavelength recordings, Fluo-3 allowed to follow, semi-quantitatively but with high time resolution, [Ca2+]i changes by rapid confocal laser scanning microscopy (CLSM). Resting values are between 50 and 70 nM in the strains analysed (7S wild type, as well as a non-discharge and a trichocyst-free mutant, nd9-28 degrees C and tl). In all strains [Ca2+]i first increases at the site of AED application, up to 10-fold above basal values, followed by a spillover into deeper cell regions. This might: (i) allow a vigorous Ca2+ flush during activation, and subsequently (ii) facilitate re-establishment of Ca2+ homeostasis within > or =20 seconds. Because of cell dislocation during vigorous trichocyst exocytosis, 7S cells could be reasonably analysed only by CLSM after Fluo-3 injection. In 7S cells cortical [Ca2+]i transients are strictly parallelled by trichocyst exocytosis, i.e. in the subsecond time range and precisely at the site of AED application. Injection of Ca2+ is a much less efficient trigger for exocytosis. Ca2+-buffer injections suggest a requirement of [Ca2+]i >1 to 10 microM for exocytosis to occur in response to AED. In conclusion, our data indicate: (i) correlation of cortical [Ca2+]i transients with exocytosis, as well as (ii) occurrence of a similar signal transduction mechanism in mutant cells where target structures may be defective or absent.
Klauke
N
N
Faculty of Biology, University of Konstanz, Germany.
Plattner
H
H
D000818:Animals; D001572:Benzofurans; D002118:Calcium / Q000378:metabolism*; D003911:Dextrans / Q000494:pharmacology*; D005456:Fluorescent Dyes; D007093:Imidazoles; D017136:Ion Transport / Q000187:drug effects; D010247:Paramecium / Q000378:metabolism*
D001572:Benzofurans; D003911:Dextrans; D005456:Fluorescent Dyes; D007093:Imidazoles; C082877:fura red; C039691:aminoethyldextran; D002118:Calcium