Abstract
The worldwide use of neonicotinoid pesticides has caused concern on account of their involvement in the decline of bee populations, which are key pollinators in most ecosystems. Here we describe a role of non-neuronal acetylcholine (ACh) for breeding of Apis mellifera carnica and a so far unknown effect of neonicotinoids on non-target insects. Royal jelly or larval food are produced by the hypopharyngeal gland of nursing bees and contain unusually high ACh concentrations (4–8 mM). ACh is extremely well conserved in royal jelly or brood food because of the acidic pH of 4.0. This condition protects ACh from degradation thus ensuring delivery of intact ACh to larvae. Raising the pH to ≥5.5 and applying cholinesterase reduced the content of ACh substantially (by 75–90%) in larval food. When this manipulated brood was tested in artificial larval breeding experiments, the survival rate was higher with food supplemented by 100% with ACh (6 mM) than with food not supplemented with ACh. ACh release from the hypopharyngeal gland and its content in brood food declined by 80%, when honeybee colonies were exposed for 4 weeks to high concentrations of the neonicotinoids clothianidin (100 parts per billion [ppb]) or thiacloprid (8,800 ppb). Under these conditions the secretory cells of the gland were markedly damaged and brood development was severely compromised. Even field-relevant low concentrations of thiacloprid (200 ppb) or clothianidin (1 and 10 ppb) reduced ACh level in the brood food and showed initial adverse effects on brood development. Our findings indicate a hitherto unknown target of neonicotinoids to induce adverse effects on non-neuronal ACh which should be considered when re-assessing the environmental risks of these compounds. To our knowledge this is a new biological mechanism, and we suggest that, in addition to their well documented neurotoxic effects, neonicotinoids may contribute to honeybee colony losses consecutive to a reduction of the ACh content in the brood food.
Highlights
As early as 1914, Ewins provided the first evidence for non-neuronal ACh synthesized in ergot fungi [1], but in the following decades the biological role of ACh was predominantly focussed on its action as a neurotransmitter operating within the nervous system
ACh is synthesized within the hypopharyngeal gland by membranebound ChAT
It has already been reported that ACh is present in royal jelly [RJ] (683–800 μg/g), the secretion product of nursing bees to feed their queen and worker brood [7]
Summary
As early as 1914, Ewins provided the first evidence for non-neuronal ACh synthesized in ergot fungi [1], but in the following decades the biological role of ACh was predominantly focussed on its action as a neurotransmitter operating within the nervous system. The terms non-neuronal ACh and non-neuronal cholinergic system describe the expression in almost all taxa, indicating an important role from the beginning of life, that is in uni- and multicellular organisms such as bacteria, algae, protists, sponges, plants as well as almost all mammalian cells [2,3,4,5,6]. While little is known about insects, ACh has been demonstrated in royal jelly, the secretion product of nursing bees for breeding [7]. This is consistent with the proliferative and so-called trophic effects of ACh mediated via muscarinic or nicotinic receptors, as has been repeatedly demonstrated elsewhere [2, 3, 6, 8,9,10]
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