Zf. Gu et al., SMOOTH-MUSCLE CELLS FROM GUINEA-PIG STOMACH POSSESS HIGH-AFFINITY GALANIN RECEPTORS THAT MEDIATE RELAXATION, The American journal of physiology, 266(5), 1994, pp. 70000839-70000845
Galanin-like immunoactivity occurs in nerves and plexi in muscle layer
s throughout gastrointestinal tract including the stomach. Galanin can
affect gastric emptying and contraction or relaxation of gastric musc
le in different species. The aim of this study was to investigate the
direct effect of galanin on dispersed gastric smooth muscle cells and
to characterize any galanin receptors that mediated any effect. Disper
sed gastric smooth muscle cells were prepared from guinea pig stomach
by collagenase digestion. Porcine galanin (p-galanin; 1 mu M) did not
stimulate contraction when present alone; however, p-galanin (1 mu M)
inhibited carbachol-induced contraction with a half-maximal effect at
7 nM. p-Galanin (1 mu M) increased cellular adenosine 3',5'-cyclic mon
ophosphate (cAMP) content by 10 s and caused a maximal increase of 80%
over basal. I-125-galanin (porcine) bound to dispersed cells in a tim
e- and temperature-dependent manner. Binding was saturable, reversible
, and specific. Binding of I-125-galanin was inhibited almost equally
by porcine and rat galanin (K-i = 6-8 nM) but was not inhibited by the
galanin-associated peptide [preprogalanin-(108-123)]. The fragment ga
lanin-(1-16) was equally potent to rat galanin; however, the fragment
galanin-(9-29) was 56-fold less potent (K-i = 370 nM). Computer analys
is demonstrated there were two binding sites for p-galanin on gastric
smooth muscle cells, a high-affinity site (K-d = 2.6 nM) with low capa
city (B-max = 175 fmol/mg protein) and a low-affinity site (Kd = 150 n
M) with large capacity (B-max = 3,611 fmol/mg protein). I-125-galanin
binding to gastric smooth muscle cell membrane was inhibited by 5'-gua
nylyl imidodiphosphate [Gpp(NH)p] in a dose-dependent manner(IC50 = 75
nM). Computer analysis demonstrated that with Gpp(NH)p the high-affin
ity binding site was lost. These results demonstrate that gastric smoo
th muscle cells possess high-affinity galanin receptors, occupation of
which causes cAMP generation and muscle relaxation. The NH2-terminus
of galanin determines the receptor affinity that is regulated by a gua
nine nucleotide-binding protein.