Early life adversity in piglets induces long‐term upregulation of the enteric cholinergic nervous system and heightened, sex‐specific secretomotor neuron responses

Abstract

Background Early life adversity (ELA) is a risk factor for the later-life onset of gastrointestinal (GI) diseases such as irritable bowel syndrome (IBS); however, the mechanisms are poorly understood. Here, we utilized a porcine model of ELA, early weaning stress (EWS), to investigate the influence of ELA on the development and function of the enteric nervous system (ENS). Methods Female and castrated male (Male-C) piglets were weaned from their sow either at 15 d of age (EWS) or 28 d of age (late weaning control; LWC). At 60 d and 170 d of age, ileal mucosa-submucosa preparations were mounted in Ussing chambers and veratridine- and corticotropin releasing factor (CRF)-evoked short circuit current (Isc) responses were recorded as indices of secretomotor neuron function. Enteric neuron numbers and the expression of select neurotransmitters and their receptors were also measured. Key Results Compared with LWC pigs, female, but not Male-C EWS pigs exhibited heightened veratridine-induced Isc responses at 60 d and 170 d of age, that were blocked with tetrodotoxin (TTX) and atropine. Ileum from EWS pigs had higher numbers of enteric neurons that were choline acetyltransferase-positive (ChAT+). Markers of increased cholinergic signaling (increased acetylcholinesterase (AchE) and down-regulated mucosal muscarinic receptor 3 gene expression were also observed in EWS pigs. Conclusions & Inferences This study demonstrated that EWS in pigs induces lasting and sex-specific hypersensitivity of secretomotor neuron function and upregulation of the cholinergic ENS. These findings may represent a mechanistic link between ELA and lifelong susceptibility to GI diseases such as IBS.

Early weaning stress induces chronic functional diarrhea, intestinal barrier defects, and increased mast cell activity in a porcine model of early life adversity

“…In line with the present findings, we have previously demonstrated that EWS pigs exhibited increased diarrheal disease in response to a live oral challenge with enterotoxigenic E. coli . (36) In addition, more recently we showed that EWS pigs exhibited a persistent, heightened sensitivity to neural-evoked secretomotor responses mediated largely by an upregulated cholinergic nervous system (28) that was more severe in EWS female pigs compared with Male-C pigs. Additional contributing factors contributing to diarrhea in EWS pigs could be the heightened excitability of intestinal mast cells.…”

“…(48) Another potential factor contributing to persistent ileal permeability in EWS females could be the increased cholinergic tone and acetylcholine release which was demonstrated in our previous study. (49) Given the female-specific response, potential estrogen effects are also a consideration as estrogen has been shown to both positively and negatively modulate intestinal barrier properties. (50–52) However, given there were no differences in intestinal permeability between LWC Male-C and LWC female pigs at the 7 (prepubertal) or 20 week (pubertal) time point, the role of postnatal estrogens, while possible, does not fit with our the present findings.…”

“…Animal protocols were performed as in our previous studies. (28) Yorkshire-duroc cross, female and Male-C piglets (castrated at 9 days of age) were weaned from their sow at 15 days of age (EWS) or 28 days of age (LWC). Weaned pigs were housed in pens with Tenderfoot® flooring (3.7 m × 2.7 m, n = 6 pigs per pen) in the same environmentally controlled room.…”

“…Tissue free of Peyer’s patches was then mounted in a 0.3cm 2 aperture on Ussing chambers (Physiologic Instruments, Inc., Sand Diego, CA or World Precision Instruments, Sarasota, FL) as described in previous studies. (28, 31) The tissue was bathed in porcine ringer’s solution on each side of the tissue. The serosal bathing solution contained 10 mM glucose that was balanced with 10 mM mannitol on the mucosal side.…”

Stress, sex, and the enteric nervous system

“…Whether comparable changes occur in peripheral tissues namely in the gastrointestinal tract (GIT) that is endowed with the enteric nervous system (ENS) exhibiting a strikingly similar repertoire of neurons, nerve fibers, glia, neurotransmitters and modulators to that in the brain, is not known. In this issue of Neurogastroenterology & Motility, Medland et al (7) using an ELA model of early weaning stress (EWS) in pigs, where newborns are weaned on day 15 instead of day 28 (normal weaning)(8), demonstrate, for the first time in a large mammalian species, a sustained modulation of ileal enteric neuron number, phenotype, neuronal enzyme activity and mucosal epithelial cell neurosecretory functions. They also highlight a key contribution of sex to those alterations.…”