On detecting stress endocrines in hamster poop

I realize I am cutting in just under the wire, but for today's rodent blogging, I'd like to discuss three things that are very near and dear to my heart: hamsters, stress endocrines, and poop.

This wonderful little specimen is a Syrian (or golden) hamster:

(Source)

There is some evidence to show that hamsters use cortisol as a primary stress endocrine, similar to humans, which would make them a better model for studying stress responses that can be extrapolated onto humans, as opposed to rats and mice, which primarily use corticosterone. There is some debate about the extent to which hamsters use coristol, and both cortisol and corticosterone are present in hamsters, so researchers are currently duking that issue out. In any case, corticosterone is a precursor to cortisol, and both are involved in the stress endocrine cascade, which I have blogged about before.

Traditionally in such hamster stress models, measurements of plasma corticosteroids were taken from the blood, and the preferred methods of getting enough blood to do so were fatal to the animal (I will spare you the gory details, but I'm sure you can imagine). You can't take multiple measurements over time on a dead animal, so in their paper, Sex differences in the excretion of fecal glucocorticoid metabolites in the Syrian hamster, Chelini et al. investigate an alternate, much less invasive way to sample stress endocrines: by looking at their poop!

Poop is easily collected without disturbance to the study animal and can be collected many times as time progresses, which is a good thing, but the metabolism and excretion of different stress hormones are different, and can differ between animals, primarily due to sex, so Chelini and colleagues designed an experiment to attempt to validate this method. To do this, they injected hamsters with adrenocorticotropic hormone (ACTH), which is the endocrine released by the anterior pituitary to tell the adrenal cortex to start making stress endocrines (more in that in this post), and a control group received a saline injection. They collected feces in four hour intervals one day before and two days after injection, then extracted (I'm really glad I'm not the person whose job it was to vortex all those poop samples) and quantified the stress endocrine metabolites present in the feces.

Fig. 1 Variation of fecal glucocorticoid metabolites (FGM) over 24 h in male (n = 20; white boxes) and female (n = 21; light gray boxes) Syrian hamsters

The first thing they found, during the 24 hours pre-ACTH injection, was that male hamsters had more fecal glucocorticoid metabolites (FGM) than females. Male baseline measurements were about four times higher than the concentrations of FGM found in females. These concentrations differed throughout the day in different ways between males and females as well.

Figs. 2 & 3 Boxplots of fecal glucocorticoid metabolites (FGM; ng/g) of male [and female] Syrian hamsters after an i.m. injection of adrenocorticotropic hormone (ACTH; dark gray boxes) or saline solution (Control; white boxes). Respective median baseline values of the ACTH (filled circle with broken lines) and control group (open circle with dotted lines) at the same time of the day are also shown. Asterisks beside the box indicate significant differences compared with the baseline levels of the same group

Compared to controls and pre-injection measurements, the experimental group that received the ACTH injection had elevated FGM in their feces after about 8 hours post-injection in both males and females. Males experienced a spike in FGM eight hours earlier (20 hours post-injection) than females, and still had elevated levels of FGM after 48 hours, whereas females were back to baseline by that time.

The successful validation of these methods is important because, since individuals can be sampled multiple times, it allows them to serve as their own controls. Differences between individuals can vary up to ten-fold in FGM concentration! Their methods also reflect the natural sexual dimorphism in these hamsters: males have larger adrenal glands and secrete more corticosteroids than females. Interestingly, the injection procedure itself did not cause a significant rise in FGM in male controls, but it did in female controls (~32 hours post-injection, around the same time as the peak FGM of experimental females). This would imply that the injection procedure is more stressful for female than male hamsters. I don't have a good idea offhand as to why that may be.

In conclusion, our results show for the first time in the Syrian hamster that adrenocortical activity can be monitored in fecal samples in a non-invasive way. Actually, the satisfactory results generated by our [methods] demonstrated clearly that changes in glucocorticoid concentrations in the blood are well reflected in changes in FGM in both sexes. The difference we observed between male and female FGM levels are in accordance with the sex difference reported by others regarding the hamster adrenal physiology. Therefore, our study provides a humane, practical, and noninvasive alternative to blood removal, and therefore a powerful tool for stress-related studies in a species where venepuncture has traditionally been regarded as problematic. This is especially important, as hamsters are used as an animal model for medical research.

I, for one, strongly approve of the development of non-invasive, non-lethal sampling methods for fuzzy things. I mean, c'mon, look at this guy:

(Source)

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Chelini, M., Otta, E., Yamakita, C., & Palme, R. (2010). Sex differences in the excretion of fecal glucocorticoid metabolites in the Syrian hamster Journal of Comparative Physiology B, 180 (6), 919-925 DOI: 10.1007/s00360-010-0467-9