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Science 339 (6124): 1202-1204

Copyright © 2013 by the American Association for the Advancement of Science

Caffeine in Floral Nectar Enhances a Pollinator's Memory of Reward

G. A. Wright1,*, D. D. Baker2, M. J. Palmer3, D. Stabler1,2, J. A. Mustard4, E. F. Power1,2, A. M. Borland2, and P. C. Stevenson5,6

1 Centre for Behaviour and Evolution, Institute of Neuroscience, Newcastle University, Newcastle upon Tyne NE1 7RU, UK.
2 School of Biology, Newcastle University, Newcastle upon Tyne NE1 7RU, UK.
3 Division of Neuroscience, Medical Research Institute, Ninewells Medical School, University of Dundee, Dundee DD1 9SY, UK.
4 School of Life Sciences, Arizona State University, Tempe, AZ 85287, USA.
5 Jodrell Laboratory, Royal Botanic Gardens, Kew, Surrey TW9 3AB, UK.
6 Natural Resources Institute, University of Greenwich, Chatham, Kent ME4 4TB, UK.

Figure 1
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Fig. 1. (A) Caffeine concentration in Coffea and Citrus spp. and a cup of instant coffee. Caffeine concentration depended on species within each genus (Coffea: Kruskal-Wallis, {chi}22 = 28.1, P < 0.001; Citrus: Kruskal-Wallis, {chi}22 = 6.98, P = 0.030); C. canephora had the highest mean concentration of all species sampled. (B) The sum of the concentration of sucrose, glucose, and fructose (total nectar sugars) depended on species (one-way analysis of variance: F5, 161 = 4.64, P < 0.001) and was greatest in Citrus maxima and hybrids (citron, lemons, clementines). [C. can., Coffea canephora, N = 34; C. lib., Coffea liberica, N = 31; C. arab., Coffea arabica, N = 27; C. par., Citrus paradisi and hybrids, Ncp = 17; C. max., Citrus maxima and hybrids, N = 5; C. sin. and C. ret., Citrus sinensis and Citrus reticulata, NCS = 7, NCR = 5 (data for these two species were pooled).] Mean responses ± SE.


Figure 2
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Fig. 2. (A) The rate of learning of bees conditioned with an odor stimulus paired with a 0.7 M sucrose reward containing caffeine. The rate of learning was slightly greater for the bees fed caffeine in reward during conditioning (logistic regression, {chi}12 = 4.85, P = 0.028). N ≥ 79 for all groups. (B) Memory recall test for odors at 10 min (white bars) or 24 hours (red bars) after bees had been trained as in (A). Bright red bars indicate that the response at 24 hours was significantly different from the control (0.7 M sucrose) (least-squares contrasts: P < 0.05); dark red bars were not significantly different. Nectar levels of caffeine are indicated by hatching. N > 79 for each group. (C) Bees fed 0.1 mM caffeine in sucrose (orange bars) were more likely to remember the conditioned odor than sucrose alone (white bars) (logistic regression, {chi}12 = 9.04, P < 0.003) at 24 hours and 72 hours after conditioning. N = 40 per group.


Figure 3
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Fig. 3. The effect of caffeine on Kenyon cells. (A and B) Example traces from a KC in intact honeybee brain recorded under voltage-clamp [(A), VH = –73 mV) and current-clamp [(B), at resting VM), showing the increase in IM and depolarization evoked by bath application of caffeine (100 μM) and subsequent reversal by the nAChR antagonist d-TC (500 μM). (C and D) Mean data showing the reversal by d-TC (500 μM) of the effect of caffeine (Caff; 100 μM) on IM [(C); N = 6, t5 = 4.03, P = 0.010; t5 = 4.07, P = 0.010] and VM [(D); N = 6, t5 = 34.1, P < 0.001; t5 = 12.0, P < 0.001]. (E and F) Comparison of the mean effects of caffeine and DPCPX on IM [(E); Caff: N = 10, t9 = 3.84, P = 0.004; DPCPX: N = 6, t5 = 4.04, P = 0.010] and VM [(F) Caff: N = 6, t5 = 34.1, P < 0.001; DPCPX: N = 6, t5 = 3.39, P = 0.019]. (G and H) Example traces [(G); rising phase shown on an expanded time scale below] and mean data [(H); rate of rise: N = 6, t5 = 2.20, P = 0.079; {tau}decay: N = 9, t8 = 3.54, P = 0.008] showing that DPCPX (100 nM) and caffeine (100 μM) slowed the decay and, in six of nine KCs, potentiated the fast component of the response evoked by exogenous ACh. (Student's paired t test used in all comparisons.) Mean responses ± SE.


Figure 4
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Fig. 4. Bees are more likely to reject sucrose solutions containing caffeine at concentrations greater than 1 mM (logistic regression, {chi}42 = 23.4, P < 0.001; for 0.7 and 1.0 M, 1 mM caffeine versus sucrose post hoc, P < 0.05; for 0.3 M, 100 mM caffeine versus sucrose post hoc, P < 0.05). Bees were less likely to drink 0.3 M sucrose (pale pink diamonds) than 0.7 M (pink circles) or 1.0 M solutions (red triangles) (logistic regression, {chi}22 = 8.69, P = 0.013). Mean responses ± SE. N0.3M = 29, N0.7M = 100, N1.0M = 20.


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