PURINERGIC STIMULATION OF RABBIT CILIATED AIRWAY EPITHELIA - CONTROL BY MULTIPLE CALCIUM SOURCES

1. Simultaneous measurements of average intracellular calcium concentr ation ([Ca2+](i)) and ciliary beat frequency (CBF) were carried out on ciliated rabbit tracheal cells in order to determine quantitatively t he role of calcium in the regulation of mucus-transporting cilia. 2. E xtracellular ATP caused a rapid increase in both [Ca2+](i) and CBF in the 0.1-1000 mu M concentration range. The rise in [Ca2+](i) levelled off to an elevated [Ca2+](i) plateau while the cilia remained in a hig h activation state. The magnitude of the rise in [Ca2+](i) and CBF as well as the value of the elevated [Ca2+](i) plateau and the value of t he sustained CBF were dependent on the concentration of ATP in the sol ution. 3. No correlation was found between the mean values of [Ca2+](i ) and CBF at rest but a sigmoidal relationship was found to exist betw een the maximal rises of these parameters following excitation with ex tracellular ATP. This sigmoidal correlation incorporated the experimen ts where [Ca2+](i) rise was induced by depletion of internal calcium s tores with thapsigargin or by entry of calcium induced by ionomycin. 4 . Extracellular ATP caused both the release of calcium from internal s tores and calcium influx from the extracellular solution. The release of calcium was identified as originating from a thapsigarin-sensitive and a thapsigargin-insensitive calcium store. It is suggested that the release of calcium from these stores induces the initial rise in CBF. 5. The sustained activation of the cilia and elevated calcium plateau were found to be the result of the extracellular ATP-induced calcium influx. This calcium influx was insensitive to the voltage-gated calci um channel inhibitors verapamil and diltiazem, but mas completely elim inated by lowering the extracellular calcium concentration to 0.1 mu M . 6. We propose that the initial jump in the CBF is mediated by the ca lcium released from a thapsigargin-insensitive calcium store adjacent Do the cilia, while the later, and longer, rise in CBF is the result o f the calcium emanating from the thapsigargin-sensitive store which is positioned further away from the cilia within the cell cytoplasm. The calcium influx that follows is responsible for sustaining the cilia a t a high level of excitation.