ROLE OF PURINERGIC RECEPTORS IN CHLORIDE SECRETION IN CACO-2 CELLS

Purinergic receptors play an important role in regulating Cl- secretio n in epithelial cells. To explore further the role of these receptors in the intestine, we utilized the human intestinal epithelial cell lin e, Caco-2, grown on permeable membrane supports and assayed for Cl- se cretion by measuring the short-circuit current (I-sc). Stimulation of I-sc by extracellular nucleotides could be detected by day 4 and incre ased by day 10 postseeding. The magnitude of stimulation of I-sc at 10 mu M in cells at day 10 was UTP > ATP > UDP much greater than 2-methy lthioadenosine 5'-triphosphate (2-MeS-ATP) = ADP on the apical side an d UTP = 2-MeS-ATP = ATP > ADP much greater than UDP on the basolateral side. Cross-desensitization studies suggested that two different rece ptors are expressed in the apical membrane, a P-2U purinoceptor and a uridine nucleotide receptor. Two different receptors are also expresse d in the basolateral membrane, a P-2U receptor and another that reacts with both 2-MeS-ATP and ADP. This latter receptor has an unusual phar macological profile, with a reactivity for 2-MeS-ATP > ADP but not for ATP. Responses to purinergic receptor agonists were inhibited by pret reatment with ,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid- acetoxymethyl ester, thapsigargin, or quinine. Thus we suggest that an increase in intracellular Ca2+ and subsequent opening of Ca2+-activat ed K+ channel play a role in increasing driving force for Cl- to exit across the apical membrane. The role of the cystic fibrosis transmembr ane conductance regulator as a Cl- exit pathway on the apical membrane was also established.