CONTROL OF CILIA IN THE BRANCHIAL BASKET OF CIONA-INTESTINALIS (ASCIDACEA)

We investigated arrest and inactivation responses of stigmatal cilia in the branchial basket of the ascidian, Ciona intestinalis. Using an improved preparation of living tissue for microscopic imaging of ciliary responses, we found that Ca-ionophore A23187 in seawater + 50 mM Ca caused actively beating cilia to assume the upright inactive posture. while A23187 in seawater + 100 mM Ca caused transient (5-10 s) stigma-wide arrests in which the cilia lie flat against the stigmatal walls. Both responses are therefore Ca dependent, but the inactive state has a lower threshold for Ca than does arrest. Membrane permeant cAMP analogues induced >40% of the quiescent cilia within a stigma to begin beating. Saponin-extracted models of stigmatal cilia were developed to study the ionic and molecular control of ciliary activity in Ciona. Extracted cilia were stimulated to beat vigorously for >45 min by ATP-containing reactivation solution (RS). Addition of 10(-5) to 10(-3) M Ca to reactivation solution caused the cilia to stand upright (inactivate), but not to arrest. The calmodulin antagonists trifluoperazine and calmidazolium (100 mM) restored active beating when included in RS + 50-100 muM Ca, thereby reversing Ca-dependent inactivation. Addition of bovine brain calmodulin to RS + 100 muM Ca did not cause arrest of reactivated cilia. RS + 100 muM cAMP + 1 mM 3-isobutyl-1-methyl-xanthine or the catalytic subunit of cAMP-dependent protein kinase increased both the pro portion and vigor of reactivated beating. Addition of 100 muM Ca to the RS + cAMP + IBMX solution caused reactivated cilia to vibrate or twitch in an upright position, suggesting that Ca and cAMP have antagonistic effects on stigmatal cilia.