Insect Control: Biological and Synthetic Agents - Index of
Insect Control: Biological and Synthetic Agents - Index of
Insect Control: Biological and Synthetic Agents - Index of
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42 2: Indoxacarb <strong>and</strong> the Sodium Channel Blocker <strong>Insect</strong>icides<br />
Figure 7 RH-5529 raised the threshold in current-clamped crayfish stretch receptors, without affecting passive membrane<br />
resistance. Records show voltage (upper) <strong>and</strong> current (lower) traces for current steps before, during, <strong>and</strong> after application <strong>of</strong><br />
10 mM RH-5529. Corresponding traces superimposed on the right show that the spike threshold was raised by RH-5529, with no<br />
change in resting potential or in the response to hyperpolarizing pulses. (Reproduced with permission from Salgado, V.L., 1990.<br />
Mode <strong>of</strong> action <strong>of</strong> insecticidal dihydropyrazoles: selective block <strong>of</strong> impulse generation in sensory nerves. Pestic. Sci. 28, 389–411;<br />
ß Society <strong>of</strong> the Chemical Industry, permission is granted by John Wiley & Sons Ltd., on behalf <strong>of</strong> the SCI.)<br />
(Salgado, 1990), but the slowly adapting stretch<br />
receptor (SASR) <strong>of</strong> the crayfish Procambarus clarkii<br />
(Wiersma et al., 1953) was selected for studies <strong>of</strong> the<br />
cellular mechanism <strong>of</strong> action, because the greater<br />
size <strong>of</strong> its neuron enables intracellular recordings<br />
at the site <strong>of</strong> spike initiation.<br />
Pyrazolines blocked the stretch receptor in crayfish<br />
with a potency similar to that in insects. In mechanoreceptors,<br />
membrane deformation resulting from<br />
elongation activates stretch-sensitive ion channels<br />
that induce a so-called generator current that depolarizes<br />
the membrane <strong>of</strong> the spike initiation zone<br />
to the threshold for action potential generation. In<br />
other words, the generator current resulting from<br />
mechanosensory transduction is encoded as a spike<br />
train in the spike initiation zone. Insertion <strong>of</strong> a<br />
microelectrode into the spike-initiation zone <strong>of</strong> an<br />
SASR neuron allowed the function <strong>of</strong> this transduction<br />
<strong>and</strong> spike-encoding process to be studied.<br />
Figure 7 shows a SASR neuron studied under current<br />
clamp, in which current pulses <strong>of</strong> varying amplitude<br />
were injected to either hyperpolarize or depolarize<br />
the membrane. Such currents bypass the sensory<br />
transduction process <strong>and</strong> allow direct assessment <strong>of</strong><br />
the spike-encoding process. The voltage traces in response<br />
to the injected currents are shown in the upper<br />
row. For negative or hyperpolarizing pulses, represented<br />
as downward deflections <strong>of</strong> both current <strong>and</strong><br />
voltage, RH-5529 did not affect the membrane<br />
response, indicating that it did not affect passive<br />
membrane properties (Figure 7). The singular effect<br />
<strong>of</strong> RH-5529 was to raise the threshold for spike generation<br />
in response to depolarizing pulses, making it<br />
more difficult for injected currents <strong>and</strong>, by inference,<br />
for generator currents, to elicit spikes. From this result,<br />
it was immediately clear that voltage-dependent<br />
Na þ channels, whose activation determines the<br />
threshold <strong>and</strong> initiates the action potential, were<br />
blocked by the pyrazoline. This was also likely the<br />
mechanism <strong>of</strong> block in the spike initiation zones<br />
<strong>of</strong> CNS pacemaker neurons, where spikes are also<br />
generated by the activation <strong>of</strong> Na þ channels in<br />
response to depolarization by summation <strong>of</strong> synaptic<br />
inputs <strong>and</strong> pacemaker currents.<br />
2.4.4. Mechanism <strong>of</strong> Na + Channel Block<br />
In order to better underst<strong>and</strong> the action <strong>of</strong> SCBIs on<br />
Na þ channels, further studies were carried out on<br />
crayfish giant axons treated with pyrazolines, with<br />
techniques that allowed the study <strong>of</strong> Na þ channels<br />
under highly controlled conditions. A first hypothesis<br />
to explain the insensitivity <strong>of</strong> axonal Na þ channels<br />
to pyrazolines was that the compounds block<br />
Na þ channels in a voltage-dependent manner, <strong>and</strong><br />
are therefore selective for channels in the spike<br />
initiation zone. Voltage-gated Na þ channels are<br />
complex proteins, whose function is regulated by<br />
membrane potential through voltage-dependent conformational<br />
changes occurring on a timescale from<br />
less than a millisecond to several seconds (Pichon <strong>and</strong><br />
Ashcr<strong>of</strong>t, 1985). At the spike initiation zone, neurons<br />
operate near the threshold for action potential generation,<br />
in the range <strong>of</strong> 70 to 50 mV, where important<br />
conformational changes in Na þ channels occur<br />
prior to opening. In contrast, axons have a resting<br />
potential near 90 mV, where Na þ channels are predominantly<br />
in the resting state. However, when axons<br />
are depolarized to the range where Na þ channels<br />
begin to undergo conformational changes, they become<br />
sensitive to pyrazolines (Salgado, 1990). Likewise,<br />
the extracellularly recorded compound action<br />
potential from motor nerves <strong>of</strong> M. sexta abdominal<br />
ganglia, which was likewise highly insensitive to<br />
DCJW, was rendered sensitive by depolarization <strong>of</strong><br />
the nerves with a high K þ saline (Wing et al., 1998).<br />
Figure 8 shows membrane currents evoked by test<br />
pulses to 0 mV before <strong>and</strong> after equilibration with<br />
10 mM RH-3421 at various holding potentials. For<br />
each trace, after initial rapid downward <strong>and</strong> upward<br />
transients, the current became inward <strong>and</strong> peaked