Human animal interaction (HAI) studies have traditionally relied on observed behaviour to gain insights into how people and animals engage with each other. But such approaches are inevitably somewhat subjective. This leaves them open to criticism: what experimental subjects appear to be doing may not be establishing a direct causal relationship between the behaviour observed and the phenomenon being studied.
For example, if a dog appears “happy” to see its owner, is this because it really is in an elated state, or simply that it recognises the hand that feeds?
To overcome such shortcomings, HAI researchers are increasingly turning to physiological markers to try to confirm the results of behavioural studies. One way to do this is to look at hormone levels. Certain hormones have long been known to be associated with specific behaviours such as bonding and stress. If animals or people show changes in levels of these chemicals, that would provide strong evidence that observed behaviours were linked to deeper biological processes.
However, like all reputable animal researchers, HAI specialists are reluctant to carry out any procedures that might cause an animal pain or discomfort. Taking blood samples to analyse hormone levels, for instance, is far from ideal from a welfare perspective. It also alters the nature of the human-animal interaction in such a way as to fail to capture its essence. If a way could be found to establish hormone levels using more minimal invasive procedures, that would satisfy both research and welfare requirements.
That was the challenge taken up by a team at Duke University and the University of Arizona, USA as part of a WALTHAM supported research project.
They wanted to see if the levels two hormones known to play significant roles in bonding and stress responses, oxytocin and vasopressin, could be measured in dogs simply by analysing saliva samples.
Both hormones are most commonly measured in blood or urine. These are not ideal both because of the degree of invasiveness of sample collection for the former, and the problem of taking samples at precise time points for the latter. These types of measures also interfere with understanding what is actually happening during the HAI.
So the team tried out a completely new method. They used a technique called ELISA (enzyme-linked immunosorbent assays) on dog saliva. This test uses antibodies and colour change to identify a substance. ELISAs are popular across many industries and are often used as a diagnostic tool in medicine.
The results have just been published in the Journal of Neuroscience Methods.
The team collected saliva samples from specially trained dogs and tried to establish if the ELISA could accurately measure the hormones present. They found that both the hormones they were interested in were detectable by ELISA. Oxytocin concentrations in dog saliva were much higher than those typically detected in humans. Collection of salivary samples with different types of swabs, or following salivary stimulation or the consumption of food led to variance in results. The researchers also detected elevations in salivary Oxytocin during nursing.
This led to a further investigation just published in a separate paper. The research team worked with a group of Labrador retrievers and Labrador retriever × golden retriever crosses (23 females, 15 males). Half of the dogs engaged in 10 min of free-form friendly interaction with a human experimenter (HAI condition), and the other half rested quietly in the same environment, without human interaction (control condition). Blood and saliva samples were collected before, and immediately following both experimental conditions, and all samples were analysed using the ELISA test. Dogs participating in HAI exhibited a significant increase in both salivary and plasma oxytocin whereas dogs in the control group did not. Salivary vasopressin showed no change in the HAI group but increased by as much as a third in the control group. Plasma AVP decreased significantly following HAI (-13%) but did not change across time in the control condition.
“This work establishes and validates a procedure for collecting and measuring important biomarkers linked to human-animal bonding in a completely non-invasive way,” says WALTHAM’s Nancy Gee, one of the paper’s authors. “This technique will allow researchers to investigate the human-dog bond in a nearly unlimited variety of settings. This new research tool represents an important step forward for HAI researchers and for the dogs involved in the studies. However, several aspects of the findings are surprising relative to previous studies of salivary hormones in humans. This raises important questions about the biological processes through which these chemical messengers reach saliva, as well as the range of techniques used for their detection.”
Further studies will now refine the technique and look at its application to broader HAI research.