Intracellular pathways regulating ciliary beating of rat brain ependymal cells

1. The mammalian brain ventricles are lined with ciliated ependymal cells. As yet little is known about the mechanisms by which neurotransmitters regu late cilia beat frequency (CBF). 2. Application of 5-HT to ependymal cells in cultured rat brainstem slices caused CBF to increase. 5-HT had an EC50 of 30 muM and at 100 muM attained a near-maximal CBF increase of 52.7 +/- 4.1% (mean +/- S.D.) (n = 8). 3. Bathing slices in Ca2+-free solution markedly reduced the 5-HT-mediated increase in CBF: Fluorescence measurements revealed that 5-HT caused a mark ed transient elevation in cytosolic Ca2+ ([Ca2+](c)) that then slowly decre ased to a plateau level. Analysis showed that the [Ca2+](c) transient was d ue to release of Ca2+ from inositol 1,4,5-trisphosphate (IP3)-sensitive sto res; the plateau was probably due to extracellular Ca2+ influx through Ca2 release-activated Ca2+ (CRAC) channels; 4. Application of ATP caused a sustained decrease in CBF. ATP had an EC50 o f about 50 muM and 100 muM ATP resulted in a maximal 57.5 +/- 6.5%, (n = 12 ) decrease in CBF. The ATP-induced decrease in CBF was unaffected by loweri ng extracellular [Ca2+], and no changes in [Ca2+](c) were observed. Exposur e of ependymal cells to forskolin caused a decrease in CBF. Ciliated ependy mal cells loaded with caged cAMP exhibited a 54.3 +/- 7.5% (n = 9) decrease in CBF following uncaging. These results suggest that ATP I educes CBF by a Ca2+-independent cAMP-mediated pathway: 5. Application of 5-HT and adenosine-5'-O-3-thiotriphosphate (ATP-gamma -S) to acutely isolated ciliated ependymal cells resulted in CRF responses sim ilar to those of ependymal cells in cultured slices suggesting that these n eurotransmitters act directly on these cells. 6. The opposite response of ciliated ependymal cells to 5-HT and ATP provid es a novel mechanism for their active involvement in central nervous system signalling.