Effect of adenosine 3',5'-cyclic monophosphate on volume and cytoskeleton of MDCK cells

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Abstract
We examined the effect of N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (DBcAMP) on the volume and cytoskeleton of confluent cultures of Madin-Darby canine kidney (MDCK) cells. A 90-min exposure to 1 mM DBcAMP resulted in a 20% reduction in volume as measured by [14C]-urea water space. The volume in cells exposed to isobutylmethylxanthine (IBMX, 0.1 mM) was reduced by 24%. In control cultures F-actin, revealed by staining with nitrobenzoxadiazole-phallacidin, was found at the base of the cell as fibers, at the junctional region as a circumferential band, and on the apical cell surface as a mottled fluorescence. A dense pattern of microtubules, revealed by indirect immunofluorescence, was seen throughout the cell. Exposure to DBcAMP for 90 min resulted in a change of F-actin fibers into dense bundles near the periphery of the cell. This effect was even more striking when cells were exposed to IBMX. Cytochalasin B disrupted F-actin and resulted in a volume reduction similar to that in DBcAMP. Neither DBcAMP nor IBMX affected the distribution of microtubules. Moreover, colchicine, which completely disrupted the microtubules, did not change MDCK cell volume. The results suggest that DBcAMP and F-actin play a role in volume control in MDCK cells.