Responsiveness of beta-escin-permeabilized rabbit gastric gland model: effects of functional peptide fragments.
We established a beta-escin-permeabilized gland model with the use of rabbit isolated gastric glands. The glands retained an ability to secrete acid, monitored by [14C]aminopyrine accumulation, in response to cAMP, forskolin, and histamine. These responses were all inhibited by cAMP-dependent protein kinase inhibitory peptide. Myosin light-chain kinase inhibitory peptide also suppressed aminopyrine accumulation, whereas the inhibitory peptide of protein kinase C or that of calmodulin kinase II was without effect. Guanosine-5'-O-(3-thiotriphosphate) (GTPgammaS) abolished cAMP-stimulated acid secretion concomitantly, interfering with the redistribution of H+-K+-ATPase from tubulovesicles to the apical membrane. To identify the targets of GTPgammaS, effects of peptide fragments of certain GTP-binding proteins were examined. Although none of the peptides related to Rab proteins showed any effect, the inhibitory peptide of Arf protein inhibited cAMP-stimulated secretion. These results demonstrate that our new model, the beta-escin-permeabilized gland, allows the introduction of relatively large molecules, e.g., peptides, into the cell, and will be quite useful for analyzing signal transduction of parietal cell function.
Akagi
K
K
Laboratory of Pharmacology and Toxicology, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo 113-0033, Japan.
Nagao
T
T
Urushidani
T
T
D000632:Aminopyrine / Q000037:antagonists & inhibitors; D000818:Animals; D001692:Biological Transport; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004928:Escin / Q000737:chemistry / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000737:chemistry; D005744:Gastric Acid / Q000378:metabolism; D005753:Gastric Mucosa / Q000187:drug effects* / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism; D066298:In Vitro Techniques; D010446:Peptide Fragments / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D010539:Permeability; D011494:Protein Kinases / Q000737:chemistry; D011817:Rabbits
D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D010455:Peptides; D000632:Aminopyrine; C061479:protein kinase inhibitor peptide; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004928:Escin; D011494:Protein Kinases; D019204:GTP-Binding Proteins; D017506:H(+)-K(+)-Exchanging ATPase