In the U. S., cystic fibrosis is the most common lethal genetic disease in the caucasian population, affecting about 1 in 2500 live births. The genetic locus of the disease, CFTR, is an epithelial anion channel so it is not surprising that CF is characterized by abnormal chloride transport in affected epithelia. But individuals with cystic fibrosis not only have abnormal chloride transport but also have abnormal sodium transport. Certain epithelia, e.g. airway and the distal nephron, express both apical Na channels and CFTR. Activation of CFTR in these cells produces C1 reabsorption and hormones that stimulate CFTR also enhance Na reabsorption. Activation of both CFTR and epithelial Na channels (ENaC) may lead to dramatic increases in NaC1 entry into cells and associated cell swelling, so it is important that cells control NaC1 entry to avoid large changes in cell volume. We hypothesize that regulatory interactions between C1 channels and Na channels provide a mechanism by which epithelia can control net NaC1 entry across the apical membrane. Activation of CFTR is associated with an inhibition of ENaCs, providing a means by which epithelial cells can regulate net NaC1 entry. Preliminary results show that inhibition of ENaC activity is via a paracrine agent whose release is CFTR dependent.
The first aim will examine how expression and activation of CFTR alters the activity of ENaC.
This aim will determine if ATP released to the outside of cells in a CFTR-dependent manner can inhibit ENaC activity. This will be accomplished by inhibiting the extracellular action of ATP with ATP degrading enzymes; determining the rate of release of ATP from A6 monolayers; examining the coupling of purinergic receptors to ENaC inhibition; and determining if the ATP release is via CFTR-dependent exocytosis. Since modulation of net NaC1 reabsorption has important implications for cell volume regulation, we also hypothesize that ENaCs will regulate CFTR. Preliminary data show that forskolin/IBMX stimulated CFTR C1 currents are significantly increased in the presence of ENaC, suggesting that ENaCs regulate CFTR.
The second aim will examine whether ENaC-mediated regulation of CFTR is via changes in the number CFTR channels at the plasma membrane and/or changes in open probability. These studies will provide new information about mechanisms of coordinate regulation of epithelial Na and C1 transport.
