We reconstituted metachronal waves on ciliated cortical sheets prepared from detergent-extracted Paramecium multimicronucleatum cells. Ciliary movements of the cortical sheet, whose intracellular side adhered to a glass coverslip, were reactivated by perfusion of a basic reactivation medium containing ATP. In this condition, the ciliary field showed only unstable localised ripples. Addition of either cyclic AMP or cyclic GMP to the basic reactivation medium generated propagating metachronal waves characteristic of each nucleotide. In order to estimate the stability of the metachronal waves, autocorrelation coefficients were calculated from images of an 8 µm diameter region within the reactivated ciliary field. The decay time for the correlation coefficient to decrease to 0.5 was only 0.04 s in the basic reactivation medium, but was increased to 0.4 or 0.9 s by the addition of cyclic AMP or cyclic GMP, respectively. The decay time was dependent not only on the concentration of cyclic nucleotide but also on the wave frequency. In order to test whether cyclic-nucleotide-dependent phosphorylation affected the generation of waves, the ciliated cortical sheets were thiophosphorylated by incubation in ATP-gamma-S (adenosine-5'-o-3-thiotriphosphate) medium containing either cyclic AMP or cyclic GMP. Following this, perfusion with the basic reactivation medium generated metachronal waves only after cyclic GMP treatment. The effect of cyclic GMP is probably related to phosphorylation of ciliary proteins.

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