Making in vitro conditions more reflective of in vivo conditions for research on the teleost gastrointestinal tract

Abstract

To date, the majority of in vitro or ex vivo fish gastrointestinal research has been conducted under unrealistic conditions. In a living fish, ionic conditions, as well as levels of ammonia, pH, HCO3- and PCO2 differ considerably between the different regions of the gastrointestinal tract. These factors also differ from those of the saline often used in gut research. Furthermore, the oxygen gradient from the serosa to the gut lumen is rarely considered: in contrast to the serosa, the lumen is a hypoxic/anoxic environment. In addition, the gut microbiome plays a significant role in gut physiology, increasing the complexity of the in vivo gut, but replicating the microbial community for in vitro studies is exceptionally difficult. However, there are ways in which we can begin to overcome these challenges. Firstly, the luminal chemistry and PO2 in each gut compartment must be carefully considered. Secondly, although microbiological culture techniques are improving, we must learn how to maintain the microbiome diversity seen in vivo. Finally, for ex vivo studies, developing mucosal (luminal) solutions that more closely mimic the in vivo conditions will better replicate physiological processes. Within the field of mammalian gut physiology, great advances in 'gut-on-chip' devices are providing the tools to better replicate in vivo conditions; adopting and adapting this technology may assist in fish gut research initiatives. This Commentary aims to make fish gut physiologists aware of the various issues in replicating the in vivo conditions and identifies solutions as well as those areas that require further improvement.

Keywords: Chyme; Gut microbiome; Intestine; Mucosal chemistry; Stomach.

Fig. 1.

The fish gastrointestinal tract. (A) Food and water enter the mouth, and waste food and metabolites exit the anus via the rectum. The gut lumen pH, P_O2 and P_CO2, as well as the concentration of ions and ammonia, and the microbiome will vary along the gastrointestinal (GI) tract and will differ between species depending on the environment they inhabit and the food resources available. The gradients of P_O2, P_CO2 and ammonia concentration between the gut lumen and the body are influenced by the animal's internal milieu. (B) Schematic diagram of the main cells present in the gut lumen.

Fig. 2.

Conceptual illustrations of a microfluidic teleost gut-on-chip. (A) The top chamber contains the fish GI microbiome grown on a porous membrane covered in mucin, to allow the perfusion of metabolites into the middle chamber, which contains the gut epithelial cells grown on a second membrane. This forms the lumen. In the bottom (serosal) chamber, a layer of companion cells (e.g. fibroblasts) is grown on the other side of the membrane supporting the gut cells. (B) A representation of a microfluidic chip with the three separate chambers: the microbiome (grey channel), gut chamber (blue channel) and serosal chamber (pink channel). Each channel is connected to a syringe pump providing bespoke fluid to each chamber. The gut chamber is connected to a pressure chamber (yellow), which is designed to mimic the pressure changes associated with peristalsis. (C) A cross-section of the three chambers, with the pressure chamber designed to mimic the natural movement of the epithelium and a list of the chamber contents to the right. Based on the design by Marrero et al. (2021).