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1 Research Service, Department of Veterans Affairs Medical Center, Omaha, NE, USA; Pulmonary, Critical Care and Sleep Medicine, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, NE, USA
2 Pulmonary, Critical Care and Sleep Medicine, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, NE, USA
* To whom correspondence should be addressed. E-mail: twyatt{at}unmc.edu.
Previously, we have shown that the ATPase-dependent motion of cilia in bovine bronchial epithelial cells (BBEC) can be regulated through the cyclic nucleotides, cAMP via the cAMP-dependent protein kinase (PKA) and cGMP via the cGMP-dependent protein kinase (PKG). Both cyclic nucleotides cause an increase in cilia beat frequency (CBF). We hypothesize that cyclic AMP and cyclic GMP may act directly at the level of the ciliary axoneme in BBEC. To examine this, we employed a novel cell-free system utilizing detergentextracted axonemes. Axoneme movement was whole-field analyzed digitally using the Sisson Ammons Video Analysis system. A suspension of extracted axonemes remain motionless until the addition of 1mM ATP that establishes a baseline CBF similar to that seen when analyzing intact ciliated BBEC. Adding 10 µM cyclic AMP or 10 µM cyclic GMP increases CBF beyond the established ATP baseline. However, the cyclic nucleotides did not stimulate CBF in the absence of ATP. Therefore, the combination of cAMP and cGMP augment ATP-driven CBF increases at the level of isolated axoneme.
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