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Activation of cAMP‐dependent Cl − currents in guinea‐pig Paneth cells without relevant evidence for CFTR expression
Author(s) -
Tsumura Takehiko,
Hazama Akihiro,
Miyoshi Takashi,
Ueda Shunji,
Okada Yasunobu
Publication year - 1998
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1998.765bd.x
Subject(s) - cystic fibrosis transmembrane conductance regulator , forskolin , patch clamp , crypt , chemistry , intracellular , microbiology and biotechnology , protein kinase a , channel blocker , paneth cell , chloride channel , biology , medicine , endocrinology , biophysics , stimulation , kinase , biochemistry , small intestine , receptor , gene , calcium , organic chemistry
1 To determine whether Paneth cells exhibit functional expression of cAMP‐activated Cl − currents and molecular expression of the cystic fibrosis transmembrane conductance regulator (CFTR), we applied whole‐cell patch clamp and single‐cell mRNA analysis by reverse transcription (RT) followed by polymerase chain reaction (PCR) amplification to single Paneth cells in crypts isolated from the guinea‐pig small intestine. 2 Prominent activation of Cl − currents was consistently observed after stimulation with dibutyryl cAMP and forskolin or with vasoactive intestinal polypeptide (VIP). The cAMP‐activated Cl − current was inhibited by removal of intracellular ATP or administration of an inhibitor of protein kinase A. 3 Many of the biophysical and pharmacological properties of the currents were phenotypically similar to those of the CFTR Cl − channel, such as the ohmic current‐voltage relationship, the anion selectivity with a Type III sequence (Br − > Cl − > I − ≫ F − ≥ gluconate − ), I − ‐induced blockage, insensitivity to a stilbene‐derivative Cl − channel blocker, and sensitivity to a carboxylate analogue Cl − channel blocker. The sensitivity of the current to glibenclamide was, however, much weaker than that reported for the CFTR Cl − channel current. In contrast to the time independence of CFTR currents, the inward component of the Paneth cell Cl − currents exhibited inactivation kinetics. 4 Expression of CFTR mRNA could not be detected by RT‐PCR analysis in almost all single Paneth cells, although its expression was consistently detected at the whole‐crypt level. The presence of a small number of CFTR‐expressing epithelial cells, which were scattered both in villi and crypts but not at the crypt base where Paneth cells were located, was demonstrated by immunocytochemistry. 5 Taken together, it appears that guinea‐pig Paneth cells functionally express cAMP‐activated Cl − conductance without relevant evidence for molecular expression of CFTR. Functional expression of VIP receptors in the Paneth cells was also demonstrated.