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A Search for Neonicotinoid Binding Sites in the Optic Lobes of Honeybees and Blowflies

Hulcoop, Ashley (2019) MOBK01 20181
Degree Projects in Molecular Biology
Abstract
Neonicotinoid insecticides are agonists to nicotinic acetylcholine receptors (nAChRs) in the brains of insects. At sub-lethal doses these chemicals cause behavioral abnormalities in several bee species, reducing overall fitness. nAChRs expressed in the optic lobes of insects bind neonicotinoids, thus may be involved in these behavioral changes. Here we attempted without success to visualize the precise location of nicotinic cholinergic receptors together with choline acetyltransferase (ChAT) in the optic lobes of Calliphora sp. and Apis mellifera using immunohistochemistry techniques and the nAChR probe α-bungarotoxin.
Popular Abstract
Lost Pollinators: Finding Neonicotinoids in Insect Brains

Flying insects are dying off. There has been a 76 % decline in Germany alone over the last 27 years. This is a problem because insects are important to the ecosystem, our economy and even our everyday lives. Without the pollination services that bees provide, we wouldn’t be able enjoy most of the fruits that we eat. An unfortunate reality is that the population decline is partly caused by neonicotinoid pesticide use on crops that we require bees to pollinate!

These pesticides are not targeted to bees, but residues end up in pollen and nectar of crops and non-crops alike. Noting this, the European Union recently voted to place severe restrictions on three widely used... (More)
Lost Pollinators: Finding Neonicotinoids in Insect Brains

Flying insects are dying off. There has been a 76 % decline in Germany alone over the last 27 years. This is a problem because insects are important to the ecosystem, our economy and even our everyday lives. Without the pollination services that bees provide, we wouldn’t be able enjoy most of the fruits that we eat. An unfortunate reality is that the population decline is partly caused by neonicotinoid pesticide use on crops that we require bees to pollinate!

These pesticides are not targeted to bees, but residues end up in pollen and nectar of crops and non-crops alike. Noting this, the European Union recently voted to place severe restrictions on three widely used neonicotinoids based on a review of thousands of published studies. This ban may cause an increase in the use of other neonicotinoids, so it is important to study their potential effects on insects. We already know that residues cause bees to get lost on foraging trips, making it less likely for them to get home. Knowing where these chemicals act in the brain might help us see how they harm bees.

Neonicotinoids Affect the Nervous System
Neonicotinoids bind to nicotinic acetylcholine receptors (nAChRs) at synapses in the nervous system, which interferes with the normal transmitting of incoming information. Different areas of the brain are responsible for receiving and processing sensory information related to how insects experience and react to the world. The optic lobes receive visual information from the eyes related to motion, colour and form, which helps the insect navigate, and could be a target for neonicotinoids. We tried to find neonicotinoid receptors in the optic lobes of honeybees (Apis mellifera) and blowflies (Calliphora sp.) by using specific molecules that attach to them.

Experimenting with Snake Venom
Certain molecules including neonicotinoids and the commonly use snake venom, α-Bungarotoxin (α-BGT) resemble the molecule that usually binds to nAChRs. This resemblance is taken advantage of to locate these receptors in the brain, since they can bind instead. Once α-BGT binds to the receptor, a fluorophore that fluoresces when exposed to a certain wavelength of light is attached, revealing its location when viewed in a fluorescence microscope. Antibodies that recognize a specific molecule are used in a similar way. We used an antibody to find choline acetyltransferase (ChAT), a protein present just upstream of nAChRs. Seeing the receptor together with ChAT very close to each other in the optic lobes was our goal, since seeing both would give more reliable information than seeing one.

We prepared fly and bee brains into sections, exposed them to antibodies and viewed them using a confocal microscope, which looks through the section layer by layer to get a 3D image of the brain. If we succeeded, then the image would light up at the specific parts of the brain that have been labelled and we would see a distinct pattern. Unfortunately, our images were fuzzy, uniform and resembled brains without any antibodies or α-BGT, which means our experiment failed! Knowing exactly where neonicotinoids bind could help in the search for pesticides that are less harmful to pollinators.

Supervisors: Elisa Rigosi and David O’Carroll
Bachelor’s Degree Project in Molecular Biology 15 hp 2018
Functional Zoology Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Hulcoop, Ashley
supervisor
organization
course
MOBK01 20181
year
type
M2 - Bachelor Degree
subject
language
English
id
8962686
date added to LUP
2018-10-31 13:36:06
date last changed
2018-10-31 13:36:06
@misc{8962686,
  abstract     = {Neonicotinoid insecticides are agonists to nicotinic acetylcholine receptors (nAChRs) in the brains of insects. At sub-lethal doses these chemicals cause behavioral abnormalities in several bee species, reducing overall fitness. nAChRs expressed in the optic lobes of insects bind neonicotinoids, thus may be involved in these behavioral changes. Here we attempted without success to visualize the precise location of nicotinic cholinergic receptors together with choline acetyltransferase (ChAT) in the optic lobes of Calliphora sp. and Apis mellifera using immunohistochemistry techniques and the nAChR probe α-bungarotoxin.},
  author       = {Hulcoop, Ashley},
  language     = {eng},
  note         = {Student Paper},
  title        = {A Search for Neonicotinoid Binding Sites in the Optic Lobes of Honeybees and Blowflies},
  year         = {2019},
}