The nudibranch mollusk Hermissenda crassicornis is normally attracted to a test light. Three days of training consisting of 50 trials per day of light paired with a rotational stimulus led to a significant increase, lasting for days, in the animal's response latency to enter a test light. The group that received light associated with rotation was significantly different from groups subjected to nonassociative control procedures. Modifications of well-known sensory networks may be related to a behavioral change that shares several operational features with associative learning.

Three days of training consisting of trials of light paired with rotation produces a long-term modification of photopositive behavior in Hermissenda crassicornis. The behavioral modification depends on the temporal association of light and rotation. For animals that received light paired with rotation, significant increases in the spontaneous activity of type B photoreceptors were correlated with changes in photopositive behavior after training. A persistent tonic depolarization of type B photoreceptors can explain the cellular changes correlated with the long-term behavioral modification produced by the temporal association of light and rotation.

The pathway supporting the conditioned stimulus (CS) is one site of plasticity that has been studied extensively in conditioned Hermissenda. Several signal transduction pathways have been implicated in classical conditioning of this preparation, although the major emphasis has been on protein kinase C. Here we provide evidence for the activation and phosphorylation of a mitogen-activated protein kinase (MAPK) pathway by one-trial and multi-trial conditioning. A one-trial in vitro conditioning procedure consisting of light (CS) paired with the application of 5-HT results in the increased incorporation of 32PO4 into proteins detected with two-dimensional gel electrophoresis. Two of the phosphoproteins have molecular weights of 44 and 42 kDa, consistent with extracellular signal-regulated protein kinases (ERK1 and ERK2). Phosphorylation of the 44 and 42 kDa proteins by one-trial conditioning was inhibited by pretreatment with PD098059, A MEK1 (ERK-Activating kinase) inhibitor. Assays of ERK activity with brain myelin basic protein as a substrate revealed greater ERK activity for the group that received one-trial conditioning compared with an unpaired control group. Western blot analysis of phosphorylated ERK using antibodies recognizing the dually phosphorylated forms of ERK1 and ERK2 showed an increase in phosphorylation after one-trial conditioning compared with unpaired controls. The increased phosphorylation of ERK after one-trial conditioning was blocked by pretreatment with PD098059. Hermissenda that received 10 or 15 conditioning trials showed significant behavioral suppression compared with pseudo-random controls. After conditioning and behavioral testing, the conditioned animals showed significantly greater phosphorylation of ERK compared with the pseudo-random controls. These results show that the ERK-MAPK signaling pathway is activated in Pavlovian conditioning of Hermissenda.

Changes in the response of B-type photoreceptors to illumination were examined in the isolated nervous systems of Hermissenda following the conditioning procedures described in the preceding paper. Analysis of the transient peak amplitude of the depolarizing generator potential at the onset of two of the three light intensities used in the behavioral studies did not reveal an enhanced photoresponse to the light. However, when the activity of the same B-photoreceptors was examined after 5 min of continuous light, there was a significant decrease in the light-adapted discharge rate and a decreased generator potential amplitude in conditioned animals as compared to the random controls. An examination of the light adapted photoresponse in preparations where spike generation and synaptic interactions were eliminated showed that the decreased photoresponse of conditioned animals was due to factors that are intrinsic to the B-photoreceptors. These results are consistent with previous work suggesting that conditioning produces substantial adaptation effects in B-photoreceptors (Crow, T. (1982) Soc. Neurosci. Abstr. 8: 824). Since the cellular changes qualitatively follow the behavioral changes and are observed at times and light intensities that are similar to those where the suppression of phototactic behavior is expressed, phototactic suppression may be directly related to the changes in the B-photoreceptors and may not require the previously proposed complex network interactions within the eyes of Hermissenda.

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