CFTR contributes to cAMP-mediated inhibition of Na+/H+ exchange in murine jejunum

Abstract

In small intestinal epithelium of wild-type (WT) mice, increases in intracellular cAMP result in stimulation of CFTR-mediated anion secretion and inhibition of electroneutral Na + absorption (via Na+/H ÷ exchange). However, several reports indicate that anion secretion and inhibition of Na t absorption do not occur following cAMP treatment of the intestine in both cystic fibrosis (CF) patients and CFTR knockout (CFTR(-)) mice. To better understand the role of CFTR during cAMP inhibition of intestinal Na ÷ absorption, we investigated: (1) whether the anion secretory function of CFTR is required for complete inhibition of Na + absorption; and (2) whether electroneutral Na ÷ absorption in the OF intestine can be normally regulated by other physiological mechanisms, e.g. extracellular hyperosmolarity. ~Na flux studies were performed on WT and CFTR(-) murine jejunum mounted in Ussing chambers (voltage-clamped) and bathed in physiological Ringers solution. Under basal conditions (+ vehicle), wr jejunum exhibited net Na ÷ absorption at a rate of 6.1 _+ 1.3/~Eq/cm2.hr (n =6). Basal Na ÷ absorption could be completely inhibited by treatment with 100/~M 5-(N-EthyI-N-isopropyl} amiloride (EIPA), a specific inhibitor of Na÷/H + exchanger activity. Treatment of the W'i jejunum with 10/~M forskolin/100 p,M IBMX to increase intracellular cAMP also completely inhibited net Na ÷ flux in WT mice (0.4 _+ 0.5 ,~Eq/cm2.hr, n=6), but did not affect net Na + absorption in the CFTR(-) jejunum (basal = 4.8 _+ 0.9 vs. treated = 4.5 _+ 0.8/~Eq/em2.hr; n=6#). To examine the role of CFTR-mediated anion secretion, Wl jejuna were simultaneously treated with forskolin/IBMX and humetanide (100/~M serosal) to inhibit CFTR-mediated Clsecretion. Bumetanide treatment recovered 46% of net Na ÷ absorption (2.2 _+ 0.4 ~Eq/cm2.hr, n = 6). To examine whether intestinal Na + absorption can be inhibited in the absence of CFTR, net Na* absorption was measured after addition of 300 mM mannitol to the bath solutions of WT and CFTR(-) jejunua. Extracellular hyperosmolarity completely inhibited Ha + absorption in both WT (-0.2 _+ 0.7/~Eq/cm2.hr, n=6) and CFTR(-) intestine (0.1 _+ 0.2 p.Eq/cm2.hr, n =9). In conclusion, functional CFTR is required for complete inhibition of intestinal Na ÷ absorption during cAMP stimulation but not during extracellular hyperosmolarity. These findings suggest the hypothesis that complete cAMP inhibition of intestinal Na ÷ absorption may require changes in epithelial cell volume resulting from CFrR-mediated anion secretion. Supported by NIH and AHA.