# Effect of artificial insemination, ruminal incubation, and esophageal tubing on cortisol concentration in blood of lactating dairy cows

## TL;DR

This study found that common farm procedures like artificial insemination, ruminal incubation, and esophageal tubing did not significantly increase stress hormone levels in dairy cows, though restraint during these procedures might have a bigger impact.

## Contribution

The study provides new evidence that routine human interventions on dairy farms do not elevate cortisol concentrations in cows, challenging assumptions about stress from these procedures.

## Key findings

- Cortisol concentrations in plasma after interventions did not differ from concentrations before the interventions.
- Cows subjected to artificial insemination had greater cortisol concentrations than those subjected to other treatments.
- Animal restraint, rather than the interventions themselves, seems to affect cortisol concentrations more.

## Abstract

Summary: We determined the concentration of cortisol in plasma before (T1) and after (T2) animals were subjected to no intervention (NEG), artificial insemination (INS), ruminal incubation (RUM), or esophageal tubing (TUB). Blood samples from 40 lactating Holstein cows were collected from the coccygeal vessels to measure cortisol concentration. Cortisol concentrations in plasma after the interventions did not differ from those before the interventions. Cows subjected to artificial insemination had greater cortisol concentrations than cows subjected to the other treatments. No interaction existed between treatment and time. Some cows subjected to insemination in a palpation rail had elevated cortisol concentrations before the intervention. Therefore, animal restraint seems to affect cortisol concentrations in plasma more than actual human intervention. In conclusion, human interventions such as INS, RUM, and TUB did not elevate the cortisol concentrations in the plasma of lactating dairy cattle under the conditions of this study.

Summary: We determined the concentration of cortisol in plasma before (T1) and after (T2) animals were subjected to no intervention (NEG), artificial insemination (INS), ruminal incubation (RUM), or esophageal tubing (TUB). Blood samples from 40 lactating Holstein cows were collected from the coccygeal vessels to measure cortisol concentration. Cortisol concentrations in plasma after the interventions did not differ from those before the interventions. Cows subjected to artificial insemination had greater cortisol concentrations than cows subjected to the other treatments. No interaction existed between treatment and time. Some cows subjected to insemination in a palpation rail had elevated cortisol concentrations before the intervention. Therefore, animal restraint seems to affect cortisol concentrations in plasma more than actual human intervention. In conclusion, human interventions such as INS, RUM, and TUB did not elevate the cortisol concentrations in the plasma of lactating dairy cattle under the conditions of this study.

•Human interventions did not elevate the cortisol concentrations in plasma.•Cows inseminated outside pens had higher cortisol concentrations before insemination.•Animal restraint might affect cortisol concentrations more than human intervention.

Human interventions did not elevate the cortisol concentrations in plasma.

Cows inseminated outside pens had higher cortisol concentrations before insemination.

Animal restraint might affect cortisol concentrations more than human intervention.

Cortisol is a hormone associated with pain, fear, distress, or discomfort. We hypothesized that human interventions increase cortisol concentrations in dairy cow plasma. Therefore, the objective of this study was to determine the concentration of cortisol in plasma before and after animals were subjected to routine on-farm and research procedures, including artificial insemination, ruminal incubation, and esophageal tubing. This study used 40 lactating Holstein cows. Experimental treatments consisted of 4 interventions. Before any intervention, a first blood sample was collected from all cows. Following this blood sample, all cows were assigned to 1 of 4 treatments. A negative control treatment consisted of collecting a second blood sample at least 30 min after the previous one, without any human intervention. Artificial insemination was performed by the herd managers from the Virginia Tech Dairy Complex (Blacksburg, VA). A rumen incubation treatment consisted of collecting a second blood sample 30 min after opening the rumen cannula, inserting the operator's arm, and pulling ruminal contents out of the rumen for 2 min, mimicking a ruminal in situ incubation, and replacing the cannula plug. The esophageal tubing treatment consisted of collecting a second blood sample 30 min after inserting an esophageal tube into the esophagus for 2 min, mimicking a ruminal drenching procedure. Blood samples were collected from the coccygeal vessels. Cortisol concentration in plasma was measured by an independent laboratory using a chemiluminescence assay. The experiment was designed as a completely randomized design with repeated measures, where cow was the subject and the pre- and postintervention sampling were the repeated observations. The statistical model included the fixed effect of treatment, the random effect of cow, the fixed effect of time, the fixed effect of the treatment by time interaction, and the random residual error. According to the Akaike information criterion, compound symmetry was used as the covariance structure for the repeated measures. Cortisol concentrations in plasma after the interventions did not differ from the concentrations in plasma before the interventions. Cows subjected to artificial insemination had greater cortisol concentrations than cows subjected to the other treatments. No interaction existed between treatment and time. Most cows subjected to insemination in a palpation rail had elevated cortisol concentrations before the intervention. Therefore, animal restraint seems to affect cortisol concentrations in plasma more than the actual human intervention. Overall, human interventions such as artificial insemination, ruminal incubation, and esophageal tubing did not elevate the cortisol concentrations in the plasma of lactating dairy cattle under the conditions of this study.