Exploration of the relationship between laboratory stress parameters and leukocyte coping capacity in ponies exposed to repetitive handling during a research project

Stress is an umbrella term indicating increased allostatic load with the potential to reach a pathological state of the organism. In this context, cortisol concentration and neutrophil granulocytes-to-lymphocyte ratio (NEU: LYM) are expected to increase. On the functional pathway, stress reduces the oxidative burst capacity of neutrophils and therefore, the leucocyte coping capacity (LCC) quantifying this oxygen radical generation of neutrophils. Therefore, we hypothesized that in ponies subjected to repetitive handling, LCC will decrease and will be negatively correlated with cortisol concentration and NEU: LYM ratio. Thus, LCC was measured over 90 min by a portable chemiluminometer in nine university owned ponies involved in a vaccination study that included handling and sampling over 42 days. The area under the curve for LCC (AUC_LCC) and LCC: NEU (AUC_{LCC: NEU}), cortisol concentration measured by a validated EIA and NEU: LYM ratio were compared before and at days 1, 3, 28, 31 and 42 after immunization by linear mixed models. Pearson correlation coefficients were calculated between AUC_LCC, AUC_{LCC: NEU}, cortisol concentration and NEU: LYM ratio.

AUC_LCC, AUC_{LCC: NEU} and cortisol concentration showed a significant decrease. AUC_LCC was moderately positively correlated with NEU: LYM and weakly positively correlated with cortisol concentration, but no correlations were found for AUC_{LCC: NEU}. The progressive decrease of AUC_LCC, AUC_{LCC: NEU}, and cortisol concentration might indicate an increasing stress load. However, this was not supported by the time course of NEU: LYM ratio or by the correlations between these parameters.

While the slight decrease in LCC over time partially supported our hypothesis, we found no evidence of correlations among AUC_{LCC: NEU} and the other measured stress parameters. This may indicate that different stress markers reflect distinct endpoints within the physiological pathways of the stress response. Future studies should confirm these preliminary results and also incorporate ethograms to obtain a more comprehensive understanding of the stress response and to further evaluate the value of LCC as a stress marker in horses.