Effects of Cholinergic and Opioid Antagonists on In Vitro Release of Met-Enkephalin, Somatostatin and Insulin-like Growth Factor-1 by and PENK Expression in Crop, Proventriculus and Duodenum of Newly Hatched Chickens

Abstract

The gastrointestinal (GI) tract is under neural, endocrine and paracrine control. The release (basal and in the presence of either cholinergic and opioid antagonists) of Met-enkephalin, insulin-like growth factor 1 (IGF-1) and somatostatin (SRIF) was determined quantitatively in vitro using explants of the crop, proventriculus and duodenum from either day 0 or day 1 chicks. In addition, the effects of cholinergic and opioid antagonists on PENK gene expression were examined. Thus, the aim of this study was to determine the roles of cholinergic and opioid receptors in the GI tract in newly hatched chickens. Moreover, the effect of IGF-1 and Met-enkephalin on cell division in duodenal explants in vitro was determined. There was both the release of Met-enkephalin from, and PENK expression in, the explants of the crop, proventriculus and duodenum tissue. This is the first report of any neuropeptide(s) being synthesized in and/or released from the crop. In vitro release of Met-enkephalin, IGF-1 and SRIF from duodenal and proventriculus explants was influenced (p < 0.01) by either cholinergic or opioid antagonists; for instance, in the presence of atropine, decreases (p < 0.001) of 17.8% and 57.7% are seen, respectively, in Met-enkephalin release and PENK expression in crop explants from day 1 chicks. Moreover, in the presence of atropine, there were increases (p < 0.001) of 47.7% and 70.9% in IGF-1 release in proventriculus explants from, respectively, day 0 and day 1 chicks. Met-enkephalin and/or IGF-1 stimulated the cell division of duodenal explants in vitro. This is the first report of Met-enkephalin release and PENK expression in the avian crop and of the effects of cholinergic or opioid antagonists on these factors. It is also the first report of either cholinergic or opioid control of IGF-1 release in the periphery of any species. There were strong relationships (p < 0.05) between the release of Met-enkephalin and that of IGF-1 in the duodenum and between the release of SRIF and that of IGF-1 in the proventriculus. This is the first report of IGF-1 and Met-enkephalin stimulating (p < 0.001) the proliferation of duodenal cells and, together, exerting a synergist effect. It is concluded that the release of Met-enkephalin, IGF-1 and SRIF from foregut regions is under tonic cholinergic and opioid control.

Keywords: chicken; cholinergic; crop; duodenum; insulin-like growth factor-1; met-enkephalin; opioid; proventriculus; somatostatin (SRIF).

Figure 1

Effects of atropine and/or hexamethonium on the in vitro release of Met-enkephalin from explants of the crop, proventriculus and duodenum from newly hatched chicks. [For a, b, c and d, different lower-case letters indicate differences p < 0.001 for the release of Met-enkephalin from the explants of the crop (A,D), proventriculus (B,E) and duodenum (C,F) tissue from 0- (A–C) and 1- (D–F) day-old chicks].

Figure 2

Effect of naltrexone (Nal) on Met-enkephalin release from and PENK expression in explants of the crop, proventriculus and duodenum in vitro. [con.—control; for a, b, c and d; different lower-case letters indicate difference p < 0.001 for release of Met-enkephalin from explants of crop (A), proventriculus (B) and duodenum (C) tissue from young chicks; PENK expression in explants of crop (D), proventriculus (E) and duodenum (F) tissue from young chicks; PENK expression is shown relative to the controls (as 1.0)].

Figure 3

Effects of atropine (At) and/or hexamethonium (Hx) on PENK expression from explants of crop, proventriculus and duodenum from newly hatched chicks [for a, b and c, different lower-case letters indicate difference p < 0.001 for PENK expression in explants of crop (A,D), proventriculus (B,E) and duodenum (C,F) tissue from 0- (A–C) and 1- (D–F) day-old chicks]. PENK expression is shown relative to the controls (1.0).

Figure 4

Effects of atropine (At) and/or hexamethonium (Hx) on in vitro release of SRIF and IGF-1 from explants of proventriculus and duodenum from newly hatched chicks. [For a, b, c and d, different lower-case letters indicate difference p < 0.001 for release of IGF-1 from explants of proventriculus (A,C) and duodenum (B,D) tissue and for release of SRIF from explants of proventriculus (E,G) and duodenum (F,H) tissue from young chicks].

Figure 5

Effect of naltrexone on IGF-1 and SRIF release from explants of the proventriculus and duodenum in vitro. [For a, b, c and d, different lower-case letters indicate difference p < 0.001 for the release of IGF-1 from explants of proventriculus (A) and duodenum (B) tissue and for the release of SRIF from explants of proventriculus (C) and duodenum (D) tissue from young chicks].

Figure 6

Effect of IGF-1 and/or Met-enkephalin on the proliferation of duodenal cells in explants incubated in vitro. [For a, b, c and d, different letters indicate difference p < 0.001].