Insecticides
632 Lecture 6 - Application of
cellular neuroscience to a
practical problem
Cellular Neuroscience Revision
 Resting potential
 Action potential
 Channels:
 voltage
gated,
 ligand gated, ionotropic & metabotropic
 Chemical synaptic transmission
Aims of lecture
 to know problems of effective application
of insecticides
 to know the main types of insecticides
 to know their site(s) of action
 possible mechanisms of resistance
Reading Matters
 Web see http://biolpc22.york.ac.uk/632
 Book:
 Tomlin,
CD S (1997) The pesticide manual
 Papers



Narahashi T (1996) Neuronal ion channels as the target sites of
insecticides Pharmacology & Toxicology 79: 1-14
ffrench-Constant, RH et al (1998) Why are there so few resistanceassociated mutations in insecticide target genes? Phil.Trans. R. Soc.
B 353 1685-1693
Matsuda et al (2001) Neonicotinoids: insecticides acting on insect
nicotinic acetylcholine receptors Trends pharm. 11:573-580
Delivering insecticide
effectively?
 rapidity
 specificity
 to
target species
 side effects
 stability
 light
& air (oxygen)
 not too persistent
 solubility
 cheap
Main targets
 development
 ecdysis
[moulting] specific to insects
 cuticle specific to insects
 respiration
 CNS
Why Knockdown
 resting insects have low metabolic demand
 unlike
mammals
 general respiratory or muscular poisons not
so good?
 knockdown insecticides
 disable
insect quickly
 OK to kill slowly

target CNS
Main classes
 organochlorine (1940s)
 cyclodiene
 organophosphorus
 pyrethroids (1975-)
 Imidacloprid (1990s)
 phenyl pyrazoles
Organophosphorus
 example: malathion
 carbamates have similar action
 more toxic to insects
 phosphorylate acetylcholinesterase
 raises [ACh], so use atropine as antidote if
humans are poisoned
Organophosphorus
 phosphate group, with two CH3 / C2H5
and one longer side chain
 often S replaces O
malathion
Carbamates also related
 originally derived from calabar beans in W
Africa
 aldicarb LD50 5mg/kg
More toxic to insects
 Insects
 oxidase  much more
toxic
 OP
 cytochrome
P450 oxidase in
mitochondria, etc
 Vertebrates
 carboxyesterase 
non-toxic
 OP
Phosphorylate
acetylcholinesterase
 active site of enzyme has
serine - OH
 active site binds P from phosphate
 half
acetylcholine
like very long (80 min)
maloxon
Cyclodiene
 e.g. Dieldrin, Lindane
 once widely used
 like other
organochlorines, very
lipid soluble
Cyclodiene mode of
dieldrin
action
 affects GABAA which
carry Cl- currents
binds to picrotoxin
site
 not GABA site
 enhances current
 faster desensitisation

GABA induced Cl- current
Cyclodiene sensitivity
 insects are more sensitive
to GABAA insecticides
because
receptor is a pentamer
 the b-subunit binds the
insecticide
 insect homooligomer b3
receptors
 mammals have
heterooligomer a b g

Phenyl pyrazoles
 fipronil
 also
targets GABAA
receptors
 same site as
Lindane
Organochlorine
 DDT
 low solubility in water, high in lipids
 at main peak of use, Americans ate
0.18mg/day
 human
mass 80kg
 Na Channel effect
 more toxic to insects
DDT
 symptoms of
poisoning are
bursty discharges
Na current effect
 Na current is slower to end in DDT
orange bar marks stimulus
Pyrethroids
 very quick knockdown
 need an oxidase inhibitor
 photostable and effective
 30g/hectare
(1% of previous insecticides\)
Pyrethroids
 major current
insecticide
 derived from
chrysanthemum
 Na channel effect
 more toxic because of
differences in Na
sequence
 may also have other
effects ?
 typically esters of
chrysanthemic acid
typical pyrethroids ...
aromatic rings & Cl
or Br contribute to
toxicity
Deltamethrin
most toxic
 No CN
 CN next to ester bond
 hyperexcitation  hypersensitive
 convulsions
 paralysis
Na channel effect
single voltage
 Sodium current lasts
longer

Voltage clamp
 Note tail current
voltage series
control
tetramethrin
Na channel effect - ii
 Unitary sodium
current lasts longer
patch clamp
 type II open even less
often but for even
longer

more toxic because
 of differences in Na channel sequence
 rat mutant isoleucine  methionine in
intracellular loop of domain 2 (I874M)
other effects ?
 Pyrethroids have been reported to affect
 calcium
channels
 GABA, ACh, glutamate receptors
Imidacloprid
 newer nicotinic
 binds to ACh
receptor
Imidacloprid ii
stimulate nerve and record EPSP
apply carbamylcholine
Summary so far
 Na+ channels targets of DDT, pyrethroids
 AChEsterase targets of OPs
 ACh receptor target of Imidacloprid
 GABAA receptor target of cyclodienes &
fipronil
Problem of Resistance
 resistance means that the insects survive!
 some species never develop,
 e.g.
tsetse flies - few offspring
 most very quick
 e.g.
mosquitoes - rapid life, many offspring
 cross resistance, e.g. to DDT and
pyrethroids because same target is used.
 [behavioural resistance]
Resistance mechanisms
 organophosphates
 organochlorine
 cyclodiene
 pyrethroids
 see Ann Rev Entomology 2000
Organophosphates
 carboxylesterase genes
amplified

e.g. in mosquito, Culex, up to
250 x copies of gene/cell
 carboxylesterase gene mutated

higher kinetics and affinity
(Tribolium)
 detoxified by
glutathione-Stransferases (i.e.
addition of
glutathione)
Organochlorine
 DDT detoxified by
glutathione-Stransferases (i.e. addition
of glutathione)
 Na channel resistance
Cyclodiene
 target site change known as Rdl
 resistance
to dieldrin
 GABAA receptor
302  serine [or glycine]
 change affects cyclodiene, picrotoxin
binding
 and reduces
desensitisation
 alanine
Pyrethroids
 non-target resistance P450 oxidase
 more
transcription giving more expression
 leads to cross-resistance to
organophosphates & carbamates
 target resistance Na+ channel
+
Na
channel
 kdr : leucine  alanine (L1014F)
9
Musca strains
 super-kdr : methionine  threonine (M918T)
Effect on currents
M918T blocks current completely
Comparative mutations
Conclusions
 Cellular neuroscience helps understand
many insecticide actions
 binding to channel proteins
 ligand-gated
 voltage
gated
 mutation leads to resistance
 target
site
 enzymatic degradation
 Web page
 http://biolpc22.york.ac.uk/632/