Acute sublethal exposure to toxic heavy metals alters honey bee (Apis mellifera) feeding behavior.

Heavy metal toxicity is an ecological concern in regions affected by processes like mining, industry, and agriculture. At sufficiently high concentrations, heavy metals are lethal to honey bees, but little is known about how sublethal doses affect honey bees or whether they will consume contaminated food. We investigated whether honey bees reject sucrose solutions contaminated with three heavy metals - cadmium, copper, and lead - as a measure of their ability to detect the metals, and whether ingesting these metals altered the bees' sucrose sensitivity. The metals elicited three different response profiles in honey bees. Cadmium was not rejected in any of the assays, and ingesting cadmium did not alter sucrose sensitivity. Copper was rejected following antennal stimulation, but was readily consumed following proboscis stimulation. Ingestion of copper did not alter sucrose sensitivity. Lead appeared to be palatable at some concentrations and altered the bees' sensitivity to and/or valuation of sucrose following antennal stimulation or ingestion of the metal. These differences likely represent unique mechanisms for detecting each metal and the pathology of toxicity. The bees' ability to detect and consume these toxic metals highlights the risk of exposure to these elements for bees living in or near contaminated environments.

Acute exposure to selenium disrupts associative conditioning and long-term memory recall in honey bees (Apis mellifera).

A plethora of toxic compounds - including pesticides, heavy metals, and metalloids - have been detected in honey bees (Apis mellifera) and their colonies. One such compound is selenium, which bees are exposed to by consuming nectar and pollen from flowers grown in contaminated areas. Though selenium is lethal at high concentrations, sublethal exposure may also impair honey bees' ability to function normally. Examining the effect of selenium exposure on learning and memory provides a sensitive assay with which to identify sublethal effects on honey bee health and behavior. To determine whether sublethal selenium exposure causes learning and memory deficits, we used proboscis extension reflex conditioning coupled with recall tests 30min and 24h post-conditioning. We exposed forager honey bees to a single sublethal dose of selenium, and 3h later we used an olfactory conditioning assay to train the bees to discriminate between one odor associated with sucrose-reinforcement and a second unreinforced odor. Following conditioning we tested short- and long-term recall of the task. Acute exposure to as little as 1.8ng of an inorganic form of selenium (sodium selenate) before conditioning caused a reduction in behavioral performance during conditioning. And, exposure to 18ng of either an inorganic form (sodium selenate) or an organic form (methylseleno-l-cysteine) of selenium caused a reduction in the bees' performance during the long-term recall test. These concentrations of selenium are lower than those found in the nectar of plants grown in selenium-contaminated soil, indicating that even low-grade selenium toxicity produces significant learning and memory impairments. This may reduce foragers' ability to effectively gather resources for the colony or nurse bees' ability to care for and maintain a healthy colony.

A proboscis extension response protocol for investigating behavioral plasticity in insects: application to basic, biomedical, and agricultural research.

Insects modify their responses to stimuli through experience of associating those stimuli with events important for survival (e.g., food, mates, threats). There are several behavioral mechanisms through which an insect learns salient associations and relates them to these events. It is important to understand this behavioral plasticity for programs aimed toward assisting insects that are beneficial for agriculture. This understanding can also be used for discovering solutions to biomedical and agricultural problems created by insects that act as disease vectors and pests. The Proboscis Extension Response (PER) conditioning protocol was developed for honey bees (Apis mellifera) over 50 years ago to study how they perceive and learn about floral odors, which signal the nectar and pollen resources a colony needs for survival. The PER procedure provides a robust and easy-to-employ framework for studying several different ecologically relevant mechanisms of behavioral plasticity. It is easily adaptable for use with several other insect species and other behavioral reflexes. These protocols can be readily employed in conjunction with various means for monitoring neural activity in the CNS via electrophysiology or bioimaging, or for manipulating targeted neuromodulatory pathways. It is a robust assay for rapidly detecting sub-lethal effects on behavior caused by environmental stressors, toxins or pesticides. We show how the PER protocol is straightforward to implement using two procedures. One is suitable as a laboratory exercise for students or for quick assays of the effect of an experimental treatment. The other provides more thorough control of variables, which is important for studies of behavioral conditioning. We show how several measures for the behavioral response ranging from binary yes/no to more continuous variable like latency and duration of proboscis extension can be used to test hypotheses. And, we discuss some pitfalls that researchers commonly encounter when they use the procedure for the first time.