# Protocol for measuring the effects of an inhibitory signal associated with danger on honey bee dopamine levels

## TL;DR

This paper describes a protocol to measure how a danger signal affects dopamine levels in honey bees.

## Contribution

The study introduces a detailed experimental protocol linking inhibitory signals to dopamine changes in honey bees.

## Key findings

- The stop signal inhibits waggle dancing in honey bees after negative experiences.
- Brain dopamine levels can be measured following simulated hornet attacks and training.
- Dopamine levels are linked to fear-like behavior in honey bees.

## Abstract

The stop signal is produced in response to negative experiences at the food source and inhibits honey bee (Apis mellifera) waggle dancing. Here, we present a protocol for measuring the effects of an inhibitory signal associated with danger on honey bee dopamine levels. We describe steps for observing honey bee colonies, training them with artificial nectar, and simulating hornet attacks. We then detail procedures for recording waggle dancing and stop signals and measuring brain dopamine levels during different treatments.

For complete details on the use and execution of this protocol, please refer to Dong et al.1

•Setting up observation colonies and training foragers to a feeder•Steps for measuring forager responses before and after hornet attacks•Procedure for quantifying bee brain dopamine levels•Increasing brain dopamine levels and studying the effect on fear-like behavior

Setting up observation colonies and training foragers to a feeder

Steps for measuring forager responses before and after hornet attacks

Procedure for quantifying bee brain dopamine levels

Increasing brain dopamine levels and studying the effect on fear-like behavior

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

The stop signal is produced in response to negative experiences at the food source and inhibits honeybee (Apis mellifera) waggle dancing. Here, we present a protocol for measuring the effects of an inhibitory signal associated with danger on honeybee dopamine levels. We describe steps for observing honeybee colonies, training them with artificial nectar, and simulating hornet attacks. We then detail procedures for recording waggle dancing and stop signals and measuring brain dopamine levels following different treatments.