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THE COUNCIL FOR TOBACCO RESEARCH-U.S.A., INC. TO: DR. S. C. SOMMERS AND STAFF FROM: D. H. Ford RE: Site visit with Dr. M. J. Welsh, University of Iowa College of Medicine, Iowa City, 10. Site visitors: Drs. D. H. Ford and R. Bing. Grant No. 1669 entitled, "Mechanisms Controlling Ion Transport in Airway Epithelia" Goal: To understand the cellular mechanisms which regulate and coordinate the ionic permeabilities of the basolateral and apical cell membranes in airway epithelia, utilizing isolated segments of dog tracheal posterior wall epithelia in culture as the experimental model. Collaborators: Dr. J. B. Stokes (membrane transport) and Mr. J. McCann (K+ channels in airway smooth muscle) both contribute to the program. Observations: Dr. Welsh appears well funded in addition to his CTR funds, having.just received $30,000/yr from the Cystic Fibrosis Foundation as well as having NIH support for work on the Pathophysiology of the res- piratory tract. He has also recently been promoted to Associate Professor. His studies on cultured isolated sections of tracheal epithelium show that when Cl conductance increases, basolateral K+ conductance also increases. The increase in K+ conductance is secondary to the increase in C- conductance and is not associated with the Na-K-ATPase pump. Further, it does not seem to be regulated by the membrane voltage, but to be regulated by intracellular Ca. By the use of 'patch clamp' techniques, Welsh has been able to isolate single K+ channels from tultured tracheal epithelial cells. The K+ channels were activated by intracellular.Ca++ at levels of from 10-8 to 10-6M. This result suggests an active role for intracellular Ca++ in the regulation of the basolateral membrane K+ conductance. (See Fig. 1.) So-called 'secretagogues,' i.e., epinephrine, increase intra- cellular Ca-++, possibly by increasing membrane permeability. Intra- cellular Ca++ may then act to regulate ion flux by coupling the apical and basolateral cell membrane permeabilities. As discussed in our con- ference, thyroid, adrenal and estrogenic hormones may further influence ion permeability inasmuch as they depolarize excitable membranes. While Welsh and his team have preliminary evidence supporting their concept for the role of Ca++ in regulating K+ conductance, the concept is far from firmly proven.

Site Visit-Dr. M. J. Welsh Grant #1669 Page 2 Work on the exocytosis of vesicles into the apical cell membrane and subsequent recapture by endocytosis has not yet been started and hopefully will begin in the coming year. Other future investigations relate to the role of phosphorylation in the activation of K+ channels. Comment: Dr. Welsh's program is well integrated with related work ongoing in other departments in the Medical Center, particularly those involving renal transport. The group working with Welsh appear to be enthusiastic and well informed, as well as productive as indicated by several new manuscripts recently submitted (3 acknowledge CTR) as well as 5 abstracts for the various meetings this Spring (3 acknowledge CTR). They appear to be on target with their program, except for the studies on exo- and endocytosis, which they plan to be starting soon. In general, they are fulfilling the aims of the program. The study is relevant to the interests of CTR and merits continued support. DHF 4/3/85

Cell SUBMUCOSAL go; SOLUTION ouabain /3^SoIA~+.c2AL- Figure 1. According to the model, "uphill" entry of Cl at the basolateral membrane is coupled to and energized by "downhill" entry of Na. The Cl entry step probably couples the entry of two Cl, one Na, and one K. Chloride exit is passive with Cl moving down a favorable electrochemical gradient across the Cl conductive apical membrane. Sodium that enters coupled to Cl is extruded via the basolateral sodium-pump (Na-K-ATPase). K that enters in exchange for Na on the pump, as well as that which enters via the electrically neutral C1 entry step, is recycled via a K conductive basolateral membrane. In trachea, there is also electrogenic Na absorption which usually occurs at a smaller rate than that for Cl secretion. Sodium absorption is accounted for by passive Na entry via an amiloride sensitive apical conductance and exits via the Na-pump. The rate of Cl secretion is regulated by a variety of neurohumoral mediators. A fundamental feature of the model shown in figure 1 (and all transporting epithelia) is that the ion transport processes at the apical membrane are different from thcse at the basolateral membrane. This segregation of the individual ion transport processes to the two cell membranes makes it possible for an epithelium to carry out vectoral ion transport. However, the ion transport processes at the opposite sides of the cell did not function independently. There is a functional linkage between the membranes so that changes in the rate of ion transport at one membrane are coupled to changes in the rate of ion transport at the opposite membrane. In tracheal epithelium, there is a coupling of the avical membrane C1 conductance to the basolateral membrane K conductance.