Biochimica et Biophysica Acre, 1066 (1991) 115 123 © 1991 Elsevier Science Publishers B,V. 0005-2736/91/$03,50 ADONIS 00052736g100267R 115 BBAMEM 75265 Changes in cellular membrane and paracellular conductances by amphotericin B in the epithelium of the bullfrog cornea Peter S. Reinach 1,. John T. Tarvin 2 and Michel Hirseh 3 Department oj Ophthalmology, Washington Unieersity School of Medicine, St. Lords, MO (U.S.A.), z Oepartment o,f Pl~sics and Axtronomy, Murray State Uni~'ersity, Muryay, KY (U.S.A.) and .~ Centre National de la Recherche Scientif~e, Laburatuire de Micrt~copie Electroni¢tue Ap~iquee ala Bidogie, Paris fFranc¢) (Received 30 July 1990) (Revised manuscript received 18 March 1991) Key words: Cornea; Epithelium; Ampholericin B; Conductance; Capacitance; Impedance analysis; Cell oauplinll In the iselated bullfrog cornea, measurements of DE electrical parameters in conjunctien with AC impt~nce and ultrastructural analyses were used to determine the effects of 10 -s M anapheterkin B on epithelial ~ellular membrane and paracellular cendnctances. In NaC! Ringers, amphetericin B elicited a 3..~fold increase in the specific apical membrane conductance (G~/Ca); where G. and C a are the apical membrane ¢enductan~ and capacitance, respectively. The Imsolnteral membrane conductance (G b) and the ~ 1 membcaae calmdtance (C b) fell by 57% and ~ respectively. In the paracellnlar pathway, the fll~t jmtctiomd complex (Gj) wls unchanged whereas the |atecal intercellular space resistance (Rp) decreased by .f~%. The declines in Gu aml Ct, were suggestive of cell volume shrinkage because these changes were cNsistent ~ a pcevimmly described decline in intracellular K + content and redaction in exposed baselateral membrane area to current flew. Uitraalrnctucal analysis validated that ampboterkin B caused cell volume shrinkage because there was: (1) il~ren~¢~ tbldiltg d the basulateral membrane and waviness of the basal aspects- of the plasma membrane; (2) dilatation ef the lateral intercellular spaces. This agreement suggests that inh-aeellular activity decremed fullmCmB expmmt to a m ~ tin B which resulted in cell volume shrinkage and an impairment of CI - uptake ~ tl~ hlmolaterul membrane. Introduction In the isolated bullfrog cornea, more than 90% of the short-circuit current (/~) is accounted for by active transepithelial CI- transport from the stromal to the tear-side bathing solution [1]. Net translocation is a consequence of uptake through a basolateral mem- brane Na/CI symport followed by etectrodiffusion across the apical membrane into the tears. The driving force for CI- uptake is the downhill ceil directed Na- gradient maintained by the activity of the Na+/K " pump in the basolaleral membrane [2]. Elcctrodiffu- stun across the apical membrane is partially dependent on K + recycling between the stmmal-side bathing solu- tion and the cell interior [3]. * Co~'respondence (present address): P.S. R©inach, Department of Physiolog~ and Endocrinolo¢¢, Medical College of Gem'gia, Au- Ilusta, GA 30912, U.S.A. The mechanism of net ion translocation across the basolateral membrane was studied following exposure to either amphotericin B or nystatin. These quasi-iono- phores increased cation apical membrane cation per- meability, which altered intracellular ion composition, and thereby elicited increases in Na'~/K ÷ pump activ- ity as well as ratesof net Na + and K + transport [4-13]. However, unexplainedty net CI- transport decreased [7]. The mechanism accounting for this decline was not further considered because the use of DC circuit analy- sis required too many simplifying assumptions whose validity may be open to question [4,5]. Accordingly, there are no studies available in which DC circuit analysis was used to analyze the effects of amplmteri- cin B on net ion transport in NaCI Ringers. Another approach is to use AC impedance analysis which has the attribute of not requiring any as~nptio~ regard- ing the selectivity of a modulatovj agent in only modify- ing a single element in the electrical equivalent [14]. We used the measurements of DC electrical param-