Learning, using natural reinforcements, in insect preparations that permit cellular neuronal analysis.

A general paradigm is described that permits testing the ability of an arthropod to learn (by operant conditioning) to alter the position of a single leg segment in order to relate to behaviorally appropriate reinforcement. The paradigm was designed so that intracellular recording from identified neurons involved would be possible during the training of a locust or grasshopper, for which extensive neuron maps are available. As a prelude to such studies, electromyograms were made from the antagonistic muscles that move the conditioned limb, which in the present experiments was the tibia of the metathoracic leg. Negative (aversive) reinforcement was provided by a loud sound/vibration and positive (reward) reinforcement by food in the form of sugar-water or fresh-growing grass. In the aversive reinforcement experiments the sound, which reflexly caused flexion, was on continually except when the tibia of one hind leg was voluntarily placed in an electronically set position "window" displaced, in extension, away from the preferred position. In feeding experiments, food was brought automatically to the mouth by a motor-driven arm when the tibia was held within a position window set away from the preferred position in either extension or flexion. Whole or headless insects learned to turn off the sound permanently, except for sporadic brief interruptions, by tonic shifting of tibial position. Insects learned to bring food to the mouth by modifying the plateau phase of a position displacement lasting for a few minutes, that was found to occur from time to time also in controls. In aversive learning, minimum times to turn off the sound were 22 sec for the easiest position and 4 min for the most difficult. The longest time in the easiest position was 1 min 40 sec and in the most difficult 39 min; excluding measurement for individuals that did not learn. In reward learning, the minimum time in the easiest position was just under 1 min, and 12 min in the most difficult position. The longest times were about 8 hr regardless of difficulty.