AJP - Gastrointestinal and Liver Physiology, Vol 261, Issue 3 476-G484, Copyright © 1991 by American Physiological Society
Inhibition of peptidases potentiates enkephalin-stimulated contraction of gastric muscle cells
D. Menozzi, Z. F. Gu, P. N. Maton and N. W. Bunnett
Department of Physiology, University of California, San Francisco 94143-0660.
Cell surface peptidases degrade enkephalins and thereby restrict the number
of molecules available to activate receptors. The effects of peptidase
inhibitors on degradation of enkephalins and on enkephalin-stimulated
contraction of gastric smooth muscle cells were examined. Muscle cells
dispersed from the guinea pig stomach degraded [Tyr1-3H] [Leu5]enkephalin
(41.6 +/- 9.0% degradation at 60 min incubation, mean +/- SD, n = 4
animals). Amastatin (10 microM, an aminopeptidase inhibitor) inhibited
degradation by 72.1 +/- 1.5% The residual peptidase activity was inhibited
by phosphoramidon (1 microM, an endopeptidase EC 3.4.24.11 inhibitor) by
58.0 +/- 11.0%. [Tyr1-125I] [Met5]enkephalin was similarly degraded.
Phosphoramidon (1 microM) inhibited the degradation of the
aminopeptidase-resistant peptide [Tyr1-3H] [D-Ala2]-[Leu5]enkephalin by
greater than 95%. [Met5]enkephalin, incubated with cells for 30 s,
stimulated contraction [50% maximal contraction (EC50) 120 +/- 50 nM, n =
6]. Pretreatment of cells with phosphoramidon alone, amastatin alone, or
phosphoramidon plus amastatin, caused 20-fold (EC50 6.5 +/- 1.1 nM), 2-fold
(EC50 63 +/- 23 nM), and 100-fold (EC50 1.1 +/- 0.3 nM) increase in potency
of [Met5]enkephalin, respectively. The results show that endopeptidase EC
3.4.24.11 and aminopeptidases contribute to degradation of enkephalins by
gastric muscle cells. The rapidity and magnitude of the potentiating
effects of the inhibitors on enkephalin-stimulated contraction suggest a
close physical relationship between the peptidases and the enkephalin
receptors.
