Behavioral and Neurological Responses to Azadirachtin and Feeding Plasticity in the Tobacco Hornworm, Manduca sexta

Phytophagous insects have evolved chemosensory systems that mediate the interface between themselves and the plant kingdom. One of these adaptations is the ability to respond to a deterrent stimulus that may indicate a plant species as noxious. Another of these adaptations is the ability to habituate to a deterrent stimulus that may be unpalatable but not toxic thereby enabling the insect to feed on an otherwise nutritious food source. Herein I focus primarily on these two aspects of insect feeding in Manduca sexta ( Lepidoptera). First I characterize the antifeedant properties of azadirachtin, a compound that is found in the Neem tree ( Azadirachta indica) and is a strong inhibitor of insect feeding. Second, I look at diet induced plastic responses of larvae to various deterrents. Feeding behavior toward neem oil extract as well as pure azadirachtin was assessed in 5 th instar larvae using two choice assays. Preference tests were set up whereby larvae could choose between consuming filter paper discs moistened with the test stimuli ( neem extract or azadirachtin) and control discs moistened with distilled water. Both test substances were found to reduce feeding in a dose dependent manner relative to water. The neem oil extract was a more powerful antifeedant than pure azadirachtin most likely due to the presence of other biologically active components. Electrophysiological recordings using pure azadirachtin were made on the medial and lateral styloconica to better understand the physiological basis of the demonstrated feeding inhibition. Electrophysiological data eliminated both the medial and lateral styloconica as the taste organs through which pure azadirachtin governs antifeedant behavior. Diet plasticity was also considered specifically by looking at the effect of prior feeding experience on the behavioral response to several deterrents. To accomplish this I carried out two separate experiments analyzing the effects of two different exposure diets. In the first experiment, larvae were exposed to a caffeine diet for 48 hours. The feeding behavior of these larvae was then evaluated and compared to larvae that had no previous exposure to caffeine. A preference test between caffeine and distilled water was carried out to measure any change in feeding behavior. The deterrence effect of caffeine was diminished upon 48 hours of exposure to a caffeine diet, a phenomenon termed habituation. I then looked to see if this effect generalizes to other deterrents. I found that habituation generalizes to some aversive stimuli but not others and I discuss the implications of such generalization. Caffeine habituated animals also appeared to be insensitive to the caffeine component of a caffeine/ KCl binary mixture, the behavioral response being similar to the response to KCL alone. The second experiment followed a similar protocol in this case looking at an exposure diet containing neem extract. Larvae were exposed to a neem diet for 48 hours and the behavioral effect of this experience was assessed in a preference test between neem extract and distilled water. In contrast to the effect that caffeine has on M. sexta, neem exposure caused an additional decrease in the preference for neem, a phenomenon I refer to as increased aversion. I looked to see if this outcome would generalize to other stimuli and found that it did in some cases and not in others. Larvae also did not show a change in preference toward pure azadirachtin after feeding on neem diet indicating that larvae were not able to habituate to the azadirachtin constituent in neem