QUICK SEARCH:   [advanced] Author: Keyword(s):
Home     Help     Feedback     Subscriptions     Archive     Search     Table of Contents

First published online March 16, 2007
Journal of Experimental Biology 210, 1225-1237 (2007)
Published by The Company of Biologists 2007
doi: 10.1242/jeb.02735

This Article Summary Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Related articles in JEB Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Sugai, R. Articles by Ito, E. Search for Related Content PubMed PubMed Citation Articles by Sugai, R. Articles by Ito, E.

One-trial conditioned taste aversion in Lymnaea: good and poor performers in long-term memory acquisition

Rio Sugai¹, Sachiyo Azami¹, Hatsuki Shiga¹, Takayuki Watanabe¹, Hisayo Sadamoto², Suguru Kobayashi², Dai Hatakeyama¹, Yutaka Fujito³, Ken Lukowiak⁴ and Etsuro Ito^1,2,*

¹ Division of Biological Sciences, Graduate School of Science, Hokkaido University, North 10, West 8, Kita-ku, Sapporo 060-0810, Japan
² Laboratory of Functional Biology, Faculty of Pharmaceutical Sciences at Kagawa Campus, Tokushima Bunri University, 1314-1 Shido, Sanuki 769-2193, Japan
³ Department of Physiology, School of Medicine, Sapporo Medical University, South 1, West 17, Chuo-ku, Sapporo 060-8556, Japan
⁴ Hotchkiss Brain Institute, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta T2N 4N1, Canada

View larger version (25K): [in this window] [in a new window]   Fig. 1. Conditioning procedures for conditioned taste aversion (CTA) in snails. (A) The one-trial conditioning procedure. (B) The 10-trial conditioning procedure. The conditioned stimulus (CS) was 5 ml of either 10 mmol l–1 sucrose solution or 0.3% carrot juice, and the unconditioned stimulus (US) in all cases was 5 ml of 10 mmol l–1 KCl solution. The CS and US were added to the Petri dish using a pipette for 15 s with a 15 s inter-stimulus interval. The inter-trial interval was 10 min in the 10-trial conditioning procedure. Before and after the conditioning procedure, the CS was applied to the lips and washed off with distilled water. Then, the feeding response was determined for 1 min as a pre-test or a post-test. A backward-conditioning (US–CS) control procedure and a naive (presented only with distilled water) control procedure were also employed. (C) Inhibitory leaning. After a 10-trial backward-conditioning procedure, a normal one-trial CTA conditioning procedure was used. As a control, the naive procedure was employed instead of the backward-conditioning procedure. We also used two different CSs, CS1 (10 mmol l–1 sucrose) and CS2 (0.3% carrot juice), and one US (10 mmol l–1 KCl). All the conditioning procedures were performed with a blind protocol.

View larger version (14K): [in this window] [in a new window]   Fig. 2. Conditioned taste aversion following a one-trial conditioning procedure in snails using sucrose as the conditioned stimulus (CS) and KCl as the unconditioned stimulus (US). (A) The ratio of good and poor performers is assessed by their response to the CS following the one-trial conditioning procedure. We defined good performers as those snails that significantly reduced their response to the CS following the one-trial conditioning procedure. This meant that good performers reduced the number of bites min–1 in response to the CS from a level of approximately 17 in the pre-test to between 0 and 1 in the post-test session. In poor performers, however, there was no significant difference in the response to the CS in the post-test session compared to the pre-test session. Thus, in each session snails responded to the CS with approximately 17 bites min–1. We found that 21 of 50 snails could be classified as good performers. That is, about 40% of snails acquired conditioned taste aversion (CTA). (B) The persistence of memory following the one-trial conditioning procedure. The numbers of snails at the pre-test were as follows: 50 for naive snails (open triangles), 50 for backward-conditioning snails (open squares), 21 for good performers (solid circles) and 29 for poor performers (open circles). All values are means ± s.e.m. The x-axis is expressed in a logarithmic scale. The difference between the feeding response of the good performers and that of the poor performers and control snails was maintained for at least 7 days at *P<0.01 (one-way ANOVA followed by the post-hoc Scheffé's test). The numbers of good performers and poor performers became 15 and 23, respectively, at 7 day because some snails withdrew their body into the shell or had died.

View larger version (10K): [in this window] [in a new window]   Fig. 3. Conditioned taste aversion (CTA) following the one-trial conditioning procedure is resistant to extinction. (A) The feeding response to the sucrose conditioned stimulus (CS) before conditioning, after conditioning and after three extinction sessions for good (open circles; N=24) and poor (solid circles; N=26) performers. (B) As in A, except that carrot juice was used as the CS. There were 21 good performers and 29 poor performers. Extinction training consisted of presentation of only the CS. The CS was presented three times with a 10-min interval after the conditioning procedure (bars). All values are means ± s.e.m. The x axes are in a logarithmic scale. The differences between the feeding response of the good performers and that of the poor performers in both cases were observed for at least 24 h at *P<0.01 (Student's t-test), showing that the memory formed by the one-trial conditioning procedure is resistant to extinction.

View larger version (9K): [in this window] [in a new window]   Fig. 4. Immediate cooling after the one-trial conditioning procedure blocks long-term memory (LTM) formation. (A) Snails (N=60) were subjected to the one-trial conditioning procedure and were then immediately (within 30 s) cooled to 4°C for 30 min. After cooling, snails were kept at room temperature and tested 1 h (N=30) and 24 h (N=30) later. There were no significant differences in the response to the conditioned stimulus (CS) following conditioning and the immediate cold block (one-way ANOVA). (B) Feeding response to the CS following the one-trial conditioning procedure after delayed cooling. Snails were cooled as in A except that the cooling was delayed until after the 10-min memory test. That is, rather than immediate cooling, cooling was delayed for 11 min. In this experiment we classified 20 snails as good performers (filled squares) and 10 (open squares) as poor performers. The bar below the graph indicates the cooling period. The x axis has a logarithmic scale. The differences between the feeding response of the good performers and that of the poor performers were observed for at least 24 h (*P<0.01, Student's t-test). All values are means ± s.e.m. These data show that the necessary new protein synthesis required for LTM is initiated within 10 min of conditioning.

View larger version (6K): [in this window] [in a new window]   Fig. 5. Food deprivation for 5 days inhibits conditioned taste aversion (CTA) memory formation. (A) The response to the conditioned stimulus (CS) in the pre- and post-test sessions following the one-trial conditioning procedure in snails deprived of food for 5 days. Memory was not formed as indicated by there being no significant difference between the pre-test and post-test response to the CS. (B) As in A, except that these snails received 10 paired backward-conditioning (CS–US) presentations. Again, long-term memory (LTM) was not observed even with this more intense conditioning. In both A and B 40 snails were used. No significant differences in the feeding response were found in either case (Student's t-test). All data are the means ± s.e.m. (N=40 each). These results show that 5 days of food deprivation alters the internal state of the snail in such a manner that memory for CTA cannot be formed.