Inactivation of the murine cftr gene abolishes cAMP-mediated but not Ca(2+)-mediated secretagogue-induced volume decrease in small-intestinal crypts.
The cellular volume of crypts isolated from 2- to 3-week-old mouse small intestine has been measured to assess the capacity of the epithelial cells to respond to secretagogues. Vasoactive intestinal polypeptide (VIP) or carbachol, respectively cAMP- and calcium-mediated secretagogues, produced a reduction crypt volume attributed to KCl loss through channels activated by the agonists. Consistent with the participation of separate chloride channels, 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) blocked the carbachol- but not the VIP-induced volume decrease, whilst glibenclamide abolished the VIP effect without affecting the carbachol-induced volume decrease. Animals homozygous for a disrupted cftr gene, introduced by gene targeting, were also used as the source for crypt isolation. In these CFTR (-/-) crypts. VIP failed to elicit any reduction in cellular volume, while the response to carbachol was indistinguishable from that seen in crypts from age-matched control animals. These results are consistent with murine CFTR being a cAMP-activated chloride channel inhibited by glibenclamide and resistant to DIDS. A separate chloride conductance activated by calcium mobilization in small-intestinal crypts appears to be independent of CFTR.
Valverde
M A
MA
AFRC Institute of Animal Physiology and Genetics Research, Cambridge Research Station, UK.
O'Brien
J A
JA
Sepúlveda
F V
FV
Ratcliff
R
R
Evans
M J
MJ
Colledge
W H
WH
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000502:physiology*; D002217:Carbachol / Q000494:pharmacology; D018118:Chloride Channels / Q000502:physiology; D000242:Cyclic AMP / Q000502:physiology*; D003550:Cystic Fibrosis / Q000235:genetics*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004553:Electric Conductivity; D005786:Gene Expression Regulation; D005905:Glyburide / Q000494:pharmacology; D007421:Intestine, Small / Q000166:cytology* / Q000187:drug effects; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D014660:Vasoactive Intestinal Peptide / Q000494:pharmacology
D018118:Chloride Channels; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D014660:Vasoactive Intestinal Peptide; D002217:Carbachol; D000242:Cyclic AMP; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D005905:Glyburide; D002118:Calcium