3.2. Gustatory responsiveness

Gustatory responsiveness, i.e. responsiveness to water and sucrose solutions of different concentrations, can be easily measured using the proboscis extension response (PER):

1. Present water and a series of sucrose concentrations (i.e. 0.1 %; 0.3 %; 1.0 %; 3.0 %; 10 %; 30 %) to the antennae of a fixed bee.

When the concentration of the sucrose solution exceeds the individual response threshold of a bee, it extends its proboscis in expectation of food (Fig. 2A).

2. Report on each stimulation whether the bee extended its proboscis or not.

3. Use the PERs to water and the different sucrose concentrations (Fig. 2B) to calculate an individual gustatory response score (GRS, Fig. 2C). This score is the sum of proboscis extension responses following a series of sucrose stimulations at the antennae. Bees with a score of "1" usually only respond to 30 % sucrose, while bees with a score of "7" respond to all sucrose stimulations with proboscis extension.

The data gained from measuring the occurrence of proboscis extension are bimodal. Either a bee extends its proboscis or it does not. Thus, frequencies of responding and non-responding animals can be used to compare different populations of bees. Alternatively, the researcher can compare the gustatory response scores of groups of bees (for review see Scheiner et al., 2003). The GRSs of a population of bees usually correlate with the gustatory response threshold of the group (Page et al., 1998), i.e. the lowest sucrose concentration that the bees can distinguish from water.

Gustatory responsiveness is an excellent indicator of the physiological state of a bee. It correlates with responsiveness to light and to odours, with division of labour and with non-associative and associative learning performance (for review see Scheiner et al., 2004). The PER assay is an inexpensive and high-throughput procedure and allows far-reaching conclusions on related behaviours such as learning and memory (Scheiner et al., 1999). It has been used frequently in neurobiological and behavioural physiological studies on honey bees (Page et al., 1998; Scheiner et al., 1999; Amdam et al., 2006; Rueppell et al., 2007). In addition, this assay can be used for examining the effects of diseases and pesticides on honey bee behaviour (Iqbal and Müller, 2007; Aliouane et al., 2009).

Fig. 2. Use of the proboscis extension response (PER) for measuring gustatory responsiveness of honey bees. A. When the antenna of a fixed bee is stimulated with a droplet of sucrose solution of sufficient concentration, the bee extends its proboscis. This reflex behaviour is termed "proboscis extension response". The number of proboscis extension responses following stimulation with a series of sucrose concentrations can be counted. This number constitutes the gustatory response score (GRS) of a bee. B. Sucrose-concentration-response curves of unresponsive bees with low gustatory response scores (GRS 0-2) and of highly responsive bees with high gustatory scores (GRS 5-7). The x-axis displays the sucrose concentration. The y-axis shows the percentage of bees extending their probosces at stimulation with that sucrose concentration. C. Gustatory response scores are a measure for the overall gustatory responsiveness and are well suited for correlation analyses. Because gustatory response scores are usually not distributed normally, median GRS (circles) and quartiles (upper and lower lines) are displayed.

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