Cyclic AMP-dependent stimulation of basolateral K(+)conductance in the rabbit conjunctival epithelium.
The regulation of Na(+)transport by cAMP in freshly isolated rabbit conjunctival epithelium, a tissue exhibiting both Cl(-)secretion and Na(+)absorption, was examined. Bulbar-palpebral segments of conjunctiva were mounted between Ussing-type hemichambers under short-circuit conditions in Cl(-)free media. In this situation, the short-circuit current (I(sc)) measures an amiloride-resistant Na(+)absorptive process in the apical-to-basolateral direction. Apical additions (each at 10 microm) of cAMP-elevating compounds, forskolin, rolipram, IBMX and epinephrine all stimulated the Na(+)-dependent I(sc)by approximately 3.5-4.5 microA cm(-2)(minimal 40% increase) and reduced transepithelial resistance (R(t)) by at least 7% (P<0.05). Pre-exposure (1 hr) to the protein kinase A (PKA) inhibitor H-89 (10 microm), which in itself inhibited the I(sc)by 0. 5 microA cm(-2), attenuated the I(sc)responses of the cAMP-elevating agents (P< 0.05, unpaired data). In reverse, H-89 promptly decreased the I(sc)by 1.5-2.5 microA cm(-2)and increased R(t)by 5% (P<0.05) in tissues pre-stimulated with either forskolin or an epinephrine plus IBMX combination. Additions of epinephrine or rolipram to apically permeabilized preparations using amphotericin B, increased the I(sc)by 12 and 22% respectively over baseline and reduced R(t)by 6% (P<0.05). Similarly, in the presence of a transepithelial K(+)gradient (apical to basolateral) and amphotericin B, cAMP elevation stimulated K diffusion across the preparation by at least 1.8 microA cm(-2)and decreased the R(t)by 4% (P<0.05), changes that were reversed by subsequent H-89 addition. Under Cl(-)rich conditions, pretreatment with 5 m m Ba(2+)reduced the basal I(sc)by 59% and blocked the cAMP-induced I(sc)stimulations typically seen in the presence of the anion. The results provide evidence for a PKA-regulated, Ba(2+)-inhibitable (voltage insensitive) basolateral K(+)conductance in rabbit conjunctival epithelial cells. The action of Cl(-)secretogogues acting via cAMP on basolateral K(+)channel activity indicates that endogenous levels of cAMP may play a role in the regulation of Cl(-)secretion and Na(+)absorption in the conjunctiva.
Turner
H C
HC
Department of Ophthalmology, Mount Sinai School of Medicine, 100th Street and 5th Avenue, New York, NY 10029, USA.
Alvarez
L J
LJ
Candia
O A
OA
D000818:Animals; D002713:Chlorine / Q000378:metabolism; D003228:Conjunctiva / Q000378:metabolism*; D046508:Culture Techniques; D000242:Cyclic AMP / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D004594:Electrophysiology; D004837:Epinephrine / Q000494:pharmacology; D004848:Epithelium / Q000378:metabolism; D005260:Female; D008297:Male; D011188:Potassium / Q000378:metabolism*; D011817:Rabbits; D012964:Sodium / Q000378:metabolism
D002713:Chlorine; D012964:Sodium; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D011188:Potassium; D004837:Epinephrine