Training of instrumental responses in dogs socially reinforced by humans

The motivational bases of the social reinforcement in human-dog relations were examined. In experiment I, performed on seven dogs, it was found that dogs were able to learn and sustain the natural responses of sitting, paw extension, and lying prostrate to conditional stimuli in the form of vocal commands reinforced only by social rewards given by the experimenter, such as petting and vocal encouragement. Overtraining did not produce deterioration of performance but, on the contrary, the continual decrease of latencies. It was evidenced that tactile stimulation plays an important role in social reward. In experiment II, instrumental responses to the auditory conditional stimuli were elaborated in two groups of dogs. The first group (nine dogs) was reinforced by food, and the second group (eight dogs) was reinforced exclusively by petting. A similar course of learning and level of performance during overtraining sessions in both groups indicated that petting serves as a good reinforcement, with rewarding value comparable to that of food reinforcement. It is suggested that a strong rewarding effect of pleasurable sensory stimuli occurs in the formation of the bond between dog and human and in the learning of different tasks.     

Limits to preference and the sensitivity of choice to rate and amount of food

Studies of choice holding food‐amount ratio constant while varying food‐rate ratio within sessions showed that local changes in preference depend on relative amount of food. The present study investigated whether sensitivity of choice to food‐rate ratio and sensitivity to food‐amount ratio are independent of one another when food‐rate ratios are varied across sessions and food‐amount ratios are varied within sessions. Food deliveries for rats’ presses on the left and right levers were scheduled according to three different food‐rate ratios of 1:1, 9:1, and 1:9; each food‐rate ratio lasted for 106 sessions and was arranged independently of seven food‐amount ratios (7:1, 6:2, 5:3, 4:4, 3:5, 2:6, and 1:7 food pellets) occurring within sessions in random sequence. Each amount ratio lasted for 10 food deliveries and was separated from another by a 60‐s blackout. Sensitivity to rate ratio was high (1.0) across food deliveries. Sensitivity to amount ratio was low when food rates were equal across alternatives, but was high when rate ratio and amount ratio opposed one another. When rate ratio and amount ratio went in the same direction, choice ratio reached an elevenfold limit which reduced sensitivity to approximately zero. We conclude that three factors affect sensitivity to amount: (1) the limit to preference, (2) the equal effect on preference of amounts greater than four pellets, and (3) the absence of differential effects of switches in amount in the equal‐rates (1:1) condition. Taken together, these findings indicate that rate and amount only sometimes combine independently as additive variables to determine preference when amount ratios vary frequently within sessions.     

DYNAMICS OF CHOICE: RELATIVE RATE AND AMOUNT AFFECT LOCAL PREFERENCE AT THREE DIFFERENT TIME SCALES

To examine extended control over local choice, the present study investigated preference in transition as food‐rate ratio provided by two levers changed across seven components within daily sessions, and food‐amount ratio changed across phases. Phase 1 arranged a food‐amount ratio of 4:1 (i.e., the left lever delivered four pellets and the right lever one pellet); Phase 2 reversed the food‐amount ratio to 1:4, and in Phase 3 the food‐amount ratio was 3:2. At a relatively extended time scale, preference was described well by a linear relation between log response ratio and log rate ratio (the generalized matching law). A small amount of carryover occurred from one rate ratio to the next but disappeared after four food deliveries. Estimates of sensitivity to food‐amount ratio were around 1.0 and were independent of rate ratio. Analysis across food deliveries within rate‐ratio components showed that the effect of a small amount was diminished by the presence of a large amount—that is, when a larger amount was present in the situation (three or four pellets), the value of a small amount (one or two pellets) became paltry. More local analysis of visits to the levers between food deliveries showed that postfood visits following a large amount were disproportionately longer than following a small amount. Continuing food deliveries from the same source tended to make visits less dependent on relative amount, but a discontinuation (i.e., food from the other lever) reinstated dependence on relative amount. Analysis at a still smaller time scale revealed preference pulses following food deliveries that confirmed the tendency toward dependence on absolute amount with continuing deliveries, and toward dependence on relative amount following discontinuations. A mathematical model based on a linear‐operator equation accounts for many of the results. The larger and longer preference following a switch to a larger amount is consistent with the idea that local preference depends on relatively extended variables even on short time scales.     

Pilot Study Investigating the Role of Therapy Dogs in Facilitating Social Interaction among Children with Autism

Ten children with autism, aged 7–10, attended 14 individual sessions of social interaction therapy. A group-comparison design was adopted. Comparing two interventions, matching and random assignment were used to form the experimental group (Animal-Assisted Play Therapy with a therapy dog) and the comparison group (an identical play therapy procedure using a doll as the dog surrogate). The within-group results using nonparametric tests showed that verbal social behavior increased significantly in the experimental group. Although the magnitude of this increase was not significantly larger than that in the comparison group, the preliminary findings suggested that the therapy dog had a positive impact on language output. The results are discussed in terms of the role that a therapy dog can fulfill as a “Speech Elicitor” for children with autism.     

Prelimbic cortex bdnf knock-down reduces instrumental responding in extinction

Anatomically selective medial prefrontal cortical projections regulate the extinction of stimulus–reinforcement associations, but the mechanisms underlying extinction of an instrumental response for reward are less well-defined and may involve structures that regulate goal-directed action. We show brain-derived neurotrophic factor (bdnf) knock-down in the prelimbic, but not orbitofrontal, cortex accelerates the initial extinction of instrumental responding for food and reduces striatal BDNF protein. When knock-down mice were provided with alternative response options to readily obtain reinforcement, extinction of the previously reinforced response was unaffected, consistent with the hypothesis that the prelimbic cortex promotes instrumental action, particularly when reinforcement is uncertain or unavailable.The rodent medial prefrontal cortex contains cytoarchitectonically distinct subregions that can be differentiated based on efferent and afferent projection patterns, with dorsal regions—including the dorsal prelimbic cortex (PLc)—sharing similar functions that differ from those of the ventromedial prefrontal cortex, which includes the medial orbitofrontal cortex (mOFC) and infralimbic cortex. These dorsal/ventral networks are considered “go” and “stop” systems, respectively, that coincidentally guide behavior (Heidbreder and Groenewegen 2003). For example, the PLc is essential for maintaining instrumental responding for food when reinforcement is uncertain (Corbit and Balleine 2003; Gourley et al. 2008a). By contrast, ventromedial structures are associated with response inhibition, particularly in the context of stimulus–reinforcement associations (Heidbreder and Groenewegen 2003).We explore the hypothesis that the PLc may also promote goal-directed responding in the absence of reinforcement, thereby slowing the extinction of a previously reinforced instrumental response. If this is the case, diminution of the biological factors essential for activity-dependent neuroplasticity and cytoskeletal structure within the PLc might be expected to shift the balance between a dorsal “go” network and ventral “stop” network. The consequence would be a rapid decline in instrumental responding during extinction training. Indeed, we report that such a manipulation—virally knocking down BDNF, which promotes long-term potentiation (Kang and Schuman 1995; Korte et al. 1995, 1996; Patterson et al. 1996) and neuronal outgrowth (McAllister et al. 1995, 1996; Xu et al. 2000a,b; Gorski et al. 2003)—within the PLc facilitates the extinction of instrumental action.In the first experiment, group-housed ≥10 wk-old male mice bred in-house and homozygous for a floxed bdnf gene (Rios et al. 2001) were anaesthetized with 1:1 2-methyl-2-butanol and tribromoethanol (Sigma Aldrich) diluted 40-fold with saline. Mice were infused into the PLc (+2.0AP, −2.8DV, ±0.1ML) with an adeno-associated virus (AAV) expressing enhanced green fluorescent protein (EGFP) ± Cre. With needles (Hamilton Co.) centered at bregma, stereotaxic coordinates were located using Kopf''s digital coordinate system with 1/100-mm resolution (David Kopf Instruments). Viral constructs were infused over 5 min with 0.5 μL/hemisphere; needles were left in place for an additional 4 min. Mice were allowed to recover for at least 2 wk, allowing for viral-mediated gene knock-down (Berton et al. 2006; Graham et al. 2007; Unger et al. 2007). All procedures were Yale University Animal Care and Use Committee approved.Mice were then food-restricted (90-min access/day) and trained to perform an instrumental response (nose poke) for food reinforcement using Med-Associates operant conditioning chambers controlled by Med-Associates software. These 25-min training sessions were conducted daily, and one, two, or three responses on one of three apertures were reinforced with a 20-mg grain-based food pellet (variable ratio 2 schedule of reinforcement; Bioserv). Two-factor (knock-down × session) analysis of variance (ANOVA) with repeated measures (RM) indicated bdnf knock-down did not affect the acquisition of instrumental responding (main effect of infusion and infusion × session interaction Fs < 1) (Fig. 1A).Open in a separate windowFigure 1.PLc bdnf knock-down decreases instrumental responding in extinction. (A) Viral-mediated PLc bdnf knock-down had no effects on the acquisition of an instrumental response for food. Responses made on the active aperture are shown (left). Responding in extinction was, however, diminished during the first extinction session (right). The break in the extinction curve represents the passage of 1 d. (B) A second group of mice was trained to respond for food before viral construct infusion. Responding during reacquisition reminder sessions after recovery was unaffected, but extinction was again immediately facilitated, as indicated by fewer responses made during sessions 1 and 2. Representative EGFP spread is inset. (C) As a control measure, this experiment was replicated in mice initially trained to perform the task, then given a mOFC, rather than PLc, bdnf knock-down. Although reinforced responding during reacquisition was diminished, responding during extinction was unchanged. Representative EGFP spread is inset. (D) In a reversal task, PLc bdnf knock-down mice did not differ in their ability to “reverse” their responding on an aperture on the opposite side of the chamber; response inhibition—extinction of responding on the previously active aperture—under these circumstances was also unchanged. (E) An enlarged EGFP image is shown (taken from inside the white box in C). EGFP radiates laterally from the infusion site, and the medial wall of the PFC can be seen at left. Symbols represent means (+ SEM) per group (*P < 0.05; ^†P = 0.07). Arrows indicate the time of knock-down, relative to testing sessions.Response extinction was then tested with 10 15-min nonreinforced sessions (five sessions/day). Here, responses made on the previously active aperture declined as expected (F_(9,72) = 6.7, P < 0.001). An interaction between group and session for responses on the active aperture was also identified (F_(9,72) = 2.3, P = 0.03). Tukey''s post-hoc tests indicated responses made during session 1 were reduced in knock-down mice (P = 0.002) (Fig. 1A). Responses made during session 2 were reduced at a trend level of significance (P = 0.07), but responding during other sessions did not differ (all Ps > 0.3), suggesting PLc bdnf knock-down facilitated initial response suppression, but not necessarily the consolidation or expression of extinction learning (Rescorla and Heth 1975).Because knock-down could conceivably regulate extinction processes via effects on initial instrumental conditioning, we trained another group of mice to perform the response prior to knock-down. Mice were then matched based on responses made during training, and surgery proceeded. After recovery, mice were given three “reacquisition” sessions identical to training sessions, during which no differences were found for responses made on the reinforced aperture (main effect of group and interaction Fs < 1) (Fig. 1B). When reinforcement was withheld, however, bdnf knock-down mice again made fewer responses relative to control mice during sessions 1 and 2 (interaction F_(9,135) = 2.3, P = 0.02; post-hoc Ps < 0.01) but not later sessions (Fig. 1B). These data further support our conclusion that PLc bdnf knock-down decreases instrumental responding during the early phases of extinction, but do not indicate whether this effect is behaviorally or anatomically specific. In this group, post-mortem EGFP distribution indicated two mice had only unilateral bdnf knock-down; these animals were excluded.To address anatomical specificity, we replicated this experiment with bdnf knocked down in the ventrally situated mOFC. This site was chosen over the infralimbic cortex because we had greater confidence we could achieve anatomically selective knock-down in this larger region. Viral constructs were infused over 3 min with 0.25 μL/hemisphere and needles aimed AP +2.3, DV −3.0, ML ±0.1 and left in place for an additional 4 min. During reacquisition, a main effect of group on responses made on the active aperture indicated mOFC bdnf knock-down, unlike PLc bdnf knock-down, decreased reinforced responding (F_(1,9) = 7.9, P = 0.02; interaction F < 1) (Fig. 1C). No effects of knock-down were, however, detected for responses made during extinction testing (group and interaction Fs < 1) (Fig. 1C). This profile is distinct from PLc bdnf knock-down mice, in which nonreinforced, but not reinforced, responding was affected. In this group, one animal with unilateral bdnf knock-down was excluded.To address behavioral specificity, mice from Figure 1B were retrained until responding for food on the active aperture was reinstated. Then, the location of the active aperture was “reversed,” such that the previously nonreinforced aperture on the opposite side of the chamber wall was reinforced. In other words, mice trained to respond on the right-side aperture were now reinforced for responding on the left-side aperture and vice versa. This “reversal” procedure allowed us to test whether PLc bdnf knock-down facilitates extinction when reinforcement is available upon the acquisition of an alternative response. We used a highly reinforcing variable ratio 2 schedule, and test sessions lasted 45 min.Under these conditions, bdnf knock-down and control mice did not differ, responding on both the previously reinforced and the newly reinforced apertures to the same degree as control mice (main effect of genotype on nonreinforced responding F_(1,14) = 1.9, P = 0.2; reinforced responding F < 1; group × session interaction F < 1) (Fig. 1D). In other words, PLc bdnf knock-down mice showed exaggerated response inhibition in the absence of reinforcement, but not when a competing response to obtain food reinforcement was available. Main effects of session on responses made on the active and inactive apertures indicated mice acquired the “reversal” (F_(3,45) = 15.2, P < 0.001; F_(3,45) = 5.7, P = 0.002, respectively).In a final behavioral experiment, male group-housed C57BL/6J mice (Charles River Laboratories, Kingston, New York), also ≥10 wk of age at the start of the experiment, were trained and infused with BDNF to evaluate whether acute PLc BDNF infusion produced the opposite effects of gene knock-down: slowed extinction. Human recombinant BDNF (Chemicon) dissolved in sterile saline in a concentration of 0.4 μg/μL (Gourley et al. 2008b) was used, with 0.2 μL/site at AP +2.0, DV −2.5, ML ±0.1 (Gourley et al. 2008a) infused over 2 min with needles left in place for 2 min after infusion.Several studies indicate BDNF has behavioral effects for several days after infusion into the striatum (Horger et al. 1999), ventral tegmental area (Lu et al. 2004), hippocampus (Shirayama et al. 2002; Gourley et al. 2008b), and prefrontal cortex (Berglind et al. 2007, 2009). Therefore, we utilized a single-infusion protocol: Food restriction resumed on day 5 after surgery, at which point mice appeared active. Testing resumed on day 7, at which point mice were subjected to three nonreinforced test sessions. bdnf knock-down mice were affected during the first and second sessions only, so this protocol would be expected to capture the window during which BDNF had effects, if any. These mice showed the typical reduction of responding across sessions (F_(2,14) = 8.6, P = 0.004) (Fig. 2). It is worth noting that responding in control mice was lower than in previous experiments; this is likely due to the more limited recovery and food restriction time after surgery. Nonetheless, we found no effect of BDNF on responding (F < 1; infusion × session interaction F_(2,14) = 1.4, P = 0.3).Open in a separate windowFigure 2.Effects of PLc BDNF microinfusion. Mice were initially trained to perform the nose poke response for food. Responses on the active aperture during training are shown at left. Mice were then infused with BDNF; subsequent instrumental responding during extinction was unaffected. (Inset) Adrenal glands were extracted and weighed after the last extinction session as a measure expected to be sensitive to PLc manipulations. Here, BDNF decreased gland weights (represented as the weight of both glands normalized to total body weight). Symbols represent means (+ SEM) per group, *P < 0.05.To verify a physiological response to PLc BDNF infusions (despite a lack of behavioral effect), we rapidly euthanized mice after the last session and extracted and weighed the adrenal glands, which secrete the hormone, corticosterone. Corticosterone secretion is sensitive to medial prefrontal cortex lesions (Diorio et al. 1993; Rangel et al. 2003) and noradrenergic depletion (Radley et al. 2008), and adrenal weights correlate with PLc BDNF expression levels (Gourley et al. 2008a). As expected, BDNF-infused mice had lighter adrenal glands (t₍₁₀₎ = 4.2, P = 0.002) (Fig. 2), indicating effects of BDNF infusion were detectable on this measure, though not on response diminution per se.Local bdnf knock-down could conceivably act in part by retarding anterograde BDNF transport to, or BDNF synthesis in, major PLc projections sites (Sobreviela et al. 1996; Altar et al. 1997; Conner et al. 1997; Kokaia et al. 1998). BDNF in those projection regions—the dorsal and ventral striatum and multiple hypothalamic subregions (Öngür and Price 2000)—as well as in the PLc itself, was therefore quantified by enzyme-linked immunosorbent assay (ELISA; Promega) in knock-down, control, and BDNF-infused mice.Brains were rapidly harvested from extinguished mice in Figures 1A and ​and2,2, and frozen and sliced into 1-mm-thick coronal sections. Brain regions were dissected bilaterally or with a single midline extraction by tissue punch (1.2-mm diameter). Tissue was then sonicated in lysis buffer (200 μL: 137 mM NaCl, 20 mM tris-Hcl [pH 8], 1% igepal, 10% glycerol, 1:100 Phosphatase Inhibitor Cocktails 1 and 2; Sigma) and stored at −80°C. ELISAs were conducted using 65 μL/sample/well and in accordance with manufacturer''s instructions. BDNF concentrations were normalized to each sample''s total protein concentration, as determined by Bradford colorimetric protein assay (Pierce). BDNF was analyzed by ANOVA or ANOVA-on-Ranks for non-normally distributed PLc values.In the PLc, BDNF was diminished in bdnf knock-down mice as expected (H_(2,18) = 0.2, P = 0.006, post-hoc Ps < 0.05), but BDNF expression in BDNF-infused mice did not differ from the control group (P > 0.05) (Fig. 3A). BDNF in the hypothalamus (F_(2,19) = 2.6, P = 0.1) and nucleus accumbens (F < 1) was not affected. By contrast, dorsal (primarily dorsomedial) striatal BDNF expression differed between groups (F_(2,20) = 5.4, P = 0.01), with knock-down mice expressing less BDNF than the BDNF-infused group (P = 0.01). BDNF in knock-down mice did not, however, significantly differ from control mice (P = 0.09).Open in a separate windowFigure 3.Quantification of BDNF in the PLc, dentate gyrus, and downstream projection sites. (A) BDNF was quantified in the PLc and major projection sites after viral-mediated gene knock-down or acute microinfusion. BDNF was diminished in the PLc of knock-down mice as expected. BDNF was also reduced in the dorsal striatum (dstri) of these animals, while other regions were unaffected by this manipulation. NAC refers to the nucleus accumbens. (B) To confirm the effects of acute BDNF infusion could be detected under some circumstances, tissue from mice infused with BDNF into the dentate gyrus (dentate) was also analyzed. Under these circumstances, elevated BDNF was detected in the hypothalamus. *P < 0.05 relative to control and BDNF-infused groups; ^§P < 0.05 relative to BDNF-infused mice; and P = 0.09 relative to control mice.For additional analyses, we conducted ELISAs on tissue from drug-naïve mice that had had a BDNF infusion of the same volume and concentration in the dorsal hippocampus, rather than PLc. As here, these animals had been subsequently tested in an instrumental conditioning task and were sacrificed 7 d after infusion (for behavioral reports, see Fig. 4 in Gourley et al. 2008b). Like the PLc, the hippocampus projects to the striatum and hypothalamus (Groenewegen et al. 1987; Kishi et al. 2000). In this instance of acute hippocampal infusion, BDNF expression was increased in hypothalamic samples (infusion × brain region interaction F_(3,27) = 3.5, P = 0.03, post-hoc P = 0.009), consistent with previous findings (Sobreviela et al. 1996). Other regions were not affected (Ps > 0.6) (Fig. 3B).Taken together, these data indicate long-term distal effects of acute BDNF infusion are detectable when BDNF is infused into the dorsal hippocampus, though not necessarily PLc. Our data do not preclude the possibility, however, that acute PLc BDNF infusion has long-term consequences for BDNF-regulated intracellular signaling cascades in these downstream sites. For example, extracellular-signal regulated kinase 1/2 phosphorylation in the nucleus accumbens is enhanced by single BDNF infusions aimed at the anterior cingulate/PLc border (Berglind et al. 2007).To summarize, we provide evidence for decreased responding in instrumentally trained mice with PLc-selective bdnf knock-down tested in extinction. Recall of extinction learning did not appear to be affected, as group differences were restricted to test sessions 1 and 2. Time of instrumental training was not a factor, as mice trained to respond for food both before and after knock-down showed a characteristically rapid decline in responding when reinforcement was withheld.Testing mice in a spatial “reversal” task, in which mice learn simultaneously to inhibit responding on one operant and respond instead on a previously nonreinforced operant, eliminated differences in nonreinforced responding between groups. In other words, in the presence of positive reinforcement, knock-down mice did not show exaggerated response inhibition. This behavioral pattern is consistent with the PLc''s role in maintaining goal-directed action particularly under low-reinforcement conditions (Corbit and Balleine 2003; Gourley et al. 2008a). If PLc bdnf played a more general role in extinction learning, one would expect PLc bdnf knock-down mice to show rapid response diminution regardless of whether reinforcement was readily available or not, but our reversal experiment clearly illustrated this was not the case.BDNF ELISA indicated the gene knock-down protocol utilized here results in an ∼48% reduction in BDNF within the PLc and a modest reduction in the downstream dorsal striatum, providing direct evidence for effects of bdnf knock-down on PLc projection neurons (though local interneurons would also be expected to have been infected). Such effects on striatal BDNF expression may be selective to chronic manipulations, as our acute infusion protocol had no consequences for expression in downstream regions, despite actions on a peripheral measure (adrenal gland weight) and evidence of downstream effects after hippocampal infusion.While we report bdnf knock-down rapidly decreased responding early in extinction, we found that acute BDNF infusion had no effects. How might we reconcile these findings? First, it is possible that prefrontal BDNF overexpression must be chronic to have behavioral effects in this task. Second, supraphysiological BDNF-induced structural destabilization and neuronal remodeling (Horch et al. 1999; Horch and Katz 2002) or activation of cortical interneurons (Rutherford et al. 1998) may have counteracted any effects on extinction. Cortical interneuron activation in particular—a process thought to stabilize cortical activity to maintain homeostasis in local circuits—could conceivably negate any effects of BDNF infusion on prefrontal projection neurons (cf., Turrigiano and Nelson 2004; see also Berglind et al. 2007). Last, while single prefrontal BDNF infusions have been reported to suppress cue-induced drug-seeking behavior (Berglind et al. 2007, 2009), such effects may be more acute and/or selectively mediated by Pavlovian, rather than instrumental, processes.Traditionally, extinction research has focused on Pavlovian fear extinction, in which the infralimbic cortex, and not PLc, is considered the major regulatory site (Quirk and Mueller 2008). Our findings suggest the PLc may, however, be indirectly involved in instrumental extinction, as bdnf knock-down facilitated rapid response diminution in the absence of reinforcement, but not when a competing response was reinforced. These findings are consistent with the idea that under normal circumstances, the PLc invigorates responding by maintaining sensitivity to reinforcement previously available upon completion of a particular instrumental action (Corbit and Balleine 2003) or previously associated with a Pavlovian cue (Vidal-Gonzalez et al. 2006). Future studies will address whether PLc BDNF is indeed critical to the maintenance of action–outcome behavior, since the mechanisms of goal-directedness are not well-characterized. This is despite the possibility that their identification may aid in therapeutically facilitating goal-directed action when response extinction is an unproductive behavioral choice.     

Agency and the Annunciation

Prior research has revealed that when healthy participants, who are not artists, are asked to draw a person who is performing an action, they are more likely to position the agent on the left and the person or object receiving this action, the patient, on the right. Thus, the goal of this study was to learn whether in works of art, such as those portraying the Annunciation of the angel Gabriel to the Blessed Virgin Mary, artists would be more likely to place the angel, who is the agent, on the left of Mary, who is the patient. We found that in our sample of 604 paintings of the Annunciation by different artists that the agent Gabriel is significantly more frequently portrayed to left of Mary. Whereas this result supports the left-agent, right-patient hypothesis, the reason for this spatial bias is not entirely known, but may be related to several factors such as the learned left to right direction of reading/writing in European languages, left-versus right-sided emotional facial expressive asymmetries, a left-sided spatial attentional bias and a spatial motor-action preference of upper extremity for making abductive (left to right) movements when using the right upper extremity. Additionally, biblical explanations and theological principles may have influenced the organization of this scene.     

The ontogenesis of lateralized behavior in the domestic cat, Felis silvestris catus

For the first time, the development of paw preferences in the domestic cat, Felis silvestris catus, is explored. Twelve cats were tested at ages 12 weeks, 6 months, and 1 year on a challenge requiring them to use one of their paws to retrieve food. To control for repeated testing of the same cats at different ages, the subjects' paw preferences were compared with those of cats tested just once, at 6 months (n = 11) or 1 year (n = 14) of age. Analysis revealed a significant effect of age on the distribution of cats' paw preferences. Cats were significantly more likely to be ambilateral than paw preferent at 12 weeks and at 6 months but more likely to display a lateral bias in paw use at 1 year. There was a significant positive correlation between cats' paw preferences at 6 months and at 1 year. Lateralized behavior was strongly sex related. Females had a greater preference for using their right paw; males were significantly more inclined to adopt their left. Analysis revealed no significant difference in the direction or strength of paw preferences of cats tested longitudinally or cross-sectionally at 6 months or 1 year of age. Findings indicate that cats develop paw preferences by 1 year and hint at a relative stability in preferred paw use over time. The strong sex effect observed strengthens the case for the influence of a biological mechanism in the emergence of motor asymmetry in cats.     

A new method for analyzing aesthetic preferences: Some theoretical considerations

The aesthetic preferences of a group of persons are obtained from their orders of sets of pictures and patterns according to “liking.” The same pictures are ordered independently by a team of experts, according to certain artistic criteria such as naturalism, composition, color, rhythm, etc. The orders of preference and orders according to the criteria are compared by correlation and matrices of correlation formed from (1) correlations between the persons' orders of preference; (2) correlations between the orders of preference and orders according to artistic criteria; and (3) correlations between the criterion orders. These matrices are symbolised byR _p ,R ₀, andR _c , respectively, and combined to form a single matrix

$$\left[ \begin{gathered} R_p R_o \hfill \\ R'_o R_c \hfill \\ \end{gathered} \right]$$     

Paw preference is not affected by postural demand in a nonprimate mammal (Felis silvestris catus)

Previously, it has been thought that handedness is unique to humans. Recently, it has been found that hand or paw preferences are common among a variety of vertebrate species. Different models have been put forth to describe the evolution of primate handedness. In this study we aimed to explore whether these models can also be used to predict manual laterality in nonprimate mammalian groups. The cat (Felis silvestris catus) is a good nonprimate model for manual laterality, as cats frequently use paws to catch and hold prey. Cats were exposed to two standardized manual laterality tasks, differing in postural demand. Subjects (N = 28) were forced to use either a stable or unstable body posture (i.e., sitting or standing vs. vertical clinging) to extract food items from a plastic box attached at two different heights. We revealed that cats exhibited paw preferences at an individual level with about 40% left, 30% right, 30% nonlateralized subjects. Postural demand was linked to task difficulty: the unstable body posture was found to be significantly more difficult than the stable body posture. However, these differences in postural demand and task difficulty did not lead to differences in direction or strength of paw preference. Findings suggested that nonprimate mammals differ from primates in their sensitivity to task related factors, such as postural demand. Results coincide with those of some prosimians, providing support for the hypothesis that postural demand and the associated task complexity became influencing factors on manual laterality in the course of primate evolution.     

The role of response-contingent incentives in lithium chloride-mediated suppression of an operant response

Three experiments investigated what role a novel incentive plays in the development of operant response suppression mediated by lithium chloride. In all experiments animals were trained to press two levers under concurrent schedules of reinforcement. In Experiment 1 responding on one lever delivered a familiar incentive (food pellets), whereas responding on an alternative lever delivered a novel incentive (sucrose solution) prior to lithium chloride injections. If lithium was administered immediately after the instrumental session, the action associated with the novel, but not with the familiar, incentive was suppressed. By comparison, in a control group for which responding on both levers led to the familiar incentive, both actions were suppressed. Experiment 2 examined whether the novelty, rather than the sensory properties, of the incentive is crucial for observing performance suppression. It was found that animals familiarized with the “target” incentive were insensitive to aversion conditioning by lithium, in that there was no difference in response rates between the action that delivered the familiar incentive from that which earned the “target”. In contrast, if animals were unfamiliar with the “target” incentive at the time of aversion conditioning, they suppressed responding on the lever that was associated with the novel incentive but did not suppress responding on the lever associated with the familiar incentive. Experiment 3 investigated the mechanism underlying instrumental performance suppression. After the completion of concurrent lever press training, novel sucrose was introduced in conjunction with the pellets for responding on one lever; responding on the other lever continued to deliver only familiar pellets. Lithium injections were then administered either immediately following the sessions or several hours after the sessions. It was found that the rate of responding on the lever associated with the contingent delivery of sucrose was suppressed below that of the pellet-alone action. By comparison, if lithium injections were administered several hours following the session, an elevation in responding on the sucrose-plus-pellet lever was observed. The outcomes of all three experiments demonstrate not only that the novelty of an incentive is important in obtaining performance suppression, but also that a novel incentive can punish instrumental responding if it has been associated with toxicosis.     

Within-Session Changes in Adjunctive and Instrumental Responding

Rats pressed levers for food delivered by several fixed interval schedules. A drinking spout or running wheel was also available during some conditions, but not during others. The rate of lever pressing, drinking and running often changed within experimental sessions. The within-session patterns of lever pressing did not differ when drinking or running was available and when it was not. The correlation between the amount of lever pressing and the amount of drinking or running at a particular time in the session was inconsistently positive or negative. Finding within-session changes in responding for adjunctive behaviors implies that the factors that produce these changes are present for both adjunctive and instrumental behavior. Finding inconsistent correlations between instrumental responding and adjunctive behaviors questions arousal and interference from adjunctive behaviors as explanations for within-session changes in instrumental responding.     

Real or imagined incomplete lateralization of function in females?

Visual field effects for 30 normal familial right-handed females were investigated with 4-point random forms known to be recognized more accurately in the right visual field by normal familial right-handed males, and the relationship between spatial ability and various performance patterns was examined. The forms were presented 2 deg in the left or right visual field. After a 10-sec delay, the subject decided whether or not a form exposed in central vision was the same as the stimulus. Incomplete lateralization of function appeared to be supported by the finding of no field differences. An alternative explanation was discussed since visual field superiority was significantly correlated with WAIS Block Design scores scaled for age, females with relatively low scores showing a right visual field superiority and females with relatively high scores showing a left visual field superiority. The experiment was repeated with 30 familial right-handed males. A significant right visual field superiority was obtained, but visual field superiorities were not correlated with measures of visuo-spatial or verbal ability.     

TEACHING CONVERSATION-RELATED SKILLS TO PREDELINQUENT GIRLS1

The use of conversation-related skills by youthful offenders can influence social interactions with adults. These behaviors are also likely to be useful to adolescents after their release from a treatment program (Journal of Applied Behavior Analysis, 1972, 5 , 343–372). Four girls, aged 13 to 15 yr, residing at Achievement Place for Girls in Lawrence, Kansas, received training on conversation-related behaviors. A multiple-baseline design across youths and across behaviors was used. Youth answer-volunteering in response to questions and three youth nonverbal components (“hand on face”, “hand at rest”, and “facial orientation”) were measured during daily 10-min sessions with a simulated guest in the group home's living room. Answer-volunteering was scored each session as the per cent of 13 “secondary” questions that the simulated guest did not have to ask following 10 “primary” questions. The three nonverbal components were scored according to their occurrence during 10-sec intervals and the resultant scores were averaged per session for an overall appropriate nonverbal score. The girls individually earned points within the home's token economy for participating in each session and additional points were awarded after training if preselected behavioral criteria were achieved for each of the two behavior categories per girl. Some of the training sessions were led by a “teaching-parent” (specially trained houseparent) while others were led by individual girls. Point consequences were administered by both the teaching-parent and by the “peer-trainers”. The average observed rate of answer-volunteering by the girls during pretraining sessions was 30% for S₁, 30% for S₂, 23% for S₃, and 68% for S₄. The average rate of answer-volunteering during posttraining sessions was: S₁ = 92%, S₂ = 89%, S₃ = 90%, and S₄ = 98%. The average nonverbal score during pretraining sessions was 82% for S₁, 53% for S₂, 60% for S₃, and 82% for S₄. The average nonverbal score during posttraining sessions was: S₁ = 98%, S₂ = 98%, S₃ = 98%, and S₄ = 100%. Videotapes of the sessions were shown in a random sequence to four adults (probation officer, social worker, etc who represented “significant others” for the youths' future success in the community. The adults judged posttraining tapes on the average as more appropriate 100% of the time for S₁, 100% of the time for S₂, 90% of the time for S₃, and 70% of the time for S₄. The study demonstrated that training of conversation-related skills is feasible with predelinquent girls, that the girls can help train each other, and that social validation of the training results is possible.     

Cultural Difference in Hand Preference: Evidence from India and Japan

Indian (N = 400) and Japanese (N = 502) adult subjects were examined for their hand preference on a 7-point scale (1 = left always, 7 = right always) of the 32-item Waterloo Handedness Questionnaire (Steenhuis & Bryden, 1989). Factor analyses of the data yielded a two-factorial structure of hand preference: skilled and unskilled. Interaction of Culture × Hand preference indicated that Indians had more right-hand preference for unskilled activities whereas Japanese subjects had more right-hand preference for skilled activities. Further analyses revealed that the frequency of middle category responses was more common in Japanese subjects. Indian subjects were found to give more extreme responses for either right- or left-hand preference. Findings are discussed in the light of cultural training given to the individuals in these two societies, which are essentially collective in nature.     

Behavior-Based Assessment of the Auditory Abilities of Brushtail Possums

Brushtail possums (Trichosurus vulpecula) were trained to press a right lever when a tone was presented (a tone‐on trial) and a left lever when a tone was not presented (a tone‐off trial) to gain access to food. During training the tone was set at 80 dB(A), with a frequency of 0.88 kH for 3 possums and of 4 kH for the other 2. Once accuracy was over 90% correct across five consecutive sessions, a test session was conducted where the intensity of the tone was reduced by 8 dB(A) over blocks of 20 trials until accuracy over a block fell below 60%. After each test session, training sessions were reintroduced and continued until accuracy was again over 90%, when another test session was conducted. This process continued until there were at least five test sessions at that tone frequency. The same procedure was then used with frequencies of 0.20, 0.88, 2, 4, 10, 12.5, 15, 20, 30, and 35 kHz. Percentage correct and d' decreased approximately linearly for all possums as tone intensity reduced. Both sets of lines were shallowest at the higher frequencies and steepest at the lower frequencies. Hit and false alarm rates mirrored each other at high frequencies but were asymmetric at lower frequencies. Equal d' contours showed that sensitivity increased from 2 to 15 kHz and continued to be high over 20 to 35 kHz. The possums remained sensitive to the 20 to 35 kHz tones even at low intensities. The present study is the first to report the abilities of possum to detect tones over this range of frequencies and its results support the findings of a microelectrode mapping survey of possums' auditory cortex.     

Comparison of contingent and noncontingent access to therapy dogs during academic tasks in children with autism spectrum disorder

This study compared contingent and noncontingent access to therapy dogs during educational tasks for children with autism spectrum disorder using a multielement design. The experimenters assessed whether initial preference for the dog predicted reinforcer efficacy and how preference changed across time. A higher response rate during contingent dog sessions than baseline sessions occurred for 4 out of 5 participants, suggesting that the dog functioned as a reinforcer. One participant engaged in a high rate of responding in both contingent and noncontingent dog conditions. Preference assessments revealed idiosyncrasies, suggesting that further research is needed into the predictive nature of initial preference assessments with animals as part of the stimulus array. The experimenters also analyzed salivary cortisol before and after sessions to determine if learning about the upcoming interaction with a dog reduced salivary cortisol in children. Cortisol was variable across participants, with only some deriving a potential physiological benefit from expecting to interact with the dog.     

Discrimination between reinforced action patterns in the rat

Laboratory rats were rewarded for face-washing, rearing, or scratching by being given the opportunity to press retractable levers for food reward. Yoked control animals received the same number of lever presentations and food rewards, but did not have to face-wash, rear, or scratch to obtain the levers. The experimental animals showed increases in number of bouts of reinforced target behavior above control levels, and the total amount of time spent face-washing increased when a 1.5-sec criterion for reinforced bout length was introduced. The activities in this experiment were made to serve a discriminative as well as an instrumental function, since the cue to tell the rat which lever to press for reward when the levers were presented was the activity that the rat had engaged in to obtain lever presentation. In two separate experiments high levels of discrimination between behaviors were obtained. Discrimination was worse following scratching than after other actions, and scratching also showed relatively poor instrumental conditioning. The relationship between Pavlovian and instrumental conditioning processes in this situation is discussed.     

The conditioned reinforcement of repeated acquisition

Three monkeys were trained to emit a chain of three responses on three separate levers in a set of six levers to obtain food. The chain producing food (correct chain) was changed each day. During a trial, a press on any lever produced a feedback stimulus; a press on a correct lever produced an additional distinctive stimulus; the third correct press produced a food pellet. Test sessions in which either the food or the distinctive stimuli were removed were interspersed with baseline sessions. In tests without food presentations, the subjects acquired the correct chain rapidly, with a level of accuracy comparable to baseline. Removing the distintive stimuli for either the first or second member of the correct chain greatly retarded acquisition of that member of the chain. Removing all distinctive stimuli often reduced accuracy throughout the chain to chance level, even though food was presented following each correct chain. These results were interpreted as evidence that the distinctive stimuli presented after correct responses functioned as conditioned reinforcers. Reductions in accuracy following an omitted distinctive stimulus indicated that they were also discriminative stimuli for correct responding in their presence.     

Covert behavior as a direct electromyographic measure of mediating responses

Electromyographic (EMG) measures (in addition to eye activity, and electroencephalograms, EEG) were taken from the tongue and arms of children under conditions of verbal oral mediation, non-oral (leg and arm) mediation based on the concepts of right and left, and no-mediation. It was found that amplitude of tongue EMG (a measure of covert oral behavior) significantly increased only for the Verbal Mediation Group, and the increase was significantly greater than for the other two (control) groups. The arms were possible loci of mediational behavior for the Non-oral Mediation Group; arm EMG was relatively large under this non-oral mediation condition, the changes being significant for the left arm. Eye movements, possible indicators of right and left mediational activity, were greater under the non-oral mediation condition, too. These psychophysiological measures of covert behavior during mediation are thus consistent with verbal mediational theory that has been based on the study of overt behavior.     

A brief retraining regulates the persistence and lability of a long-term memory

An experience extending the persistence of a memory after training Aplysia californica with inedible food also allows a consolidated memory to become sensitive to consolidation blockers. Long-term (24 h) memory is initiated by 5 min of training and is dependent on protein synthesis during the first few hours after training. By contrast, a more persistent (48 h) memory is dependent on a longer training session and on a later round of protein synthesis. When presented 24 h after training, a 3-min training that produces no memory alone can cause a memory that would have persisted for only 24 h to persist for 48 h. After a 48 h memory has been consolidated, 3 min of training also makes the memory sensitive to a protein-synthesis inhibitor. These findings suggest that a function of allowing a consolidated memory to become sensitive to blockers of protein synthesis may be to allow the memory to become more persistent.Long-term memory of an experience is dependent on a consolidation process that follows the experience. Before the memory is consolidated it is labile and can be disrupted (Lechner et al. 1999; Dudai 2004; Alberini and Taubenfeld 2008), particularly by blocking mRNA and protein synthesis (Alberini and Taubenfeld 2008; Klann and Sweatt 2008), which are needed to produce the changes in synaptic structure and function that underlie long-term memory (Sigurdsson et al. 2007; Bailey and Kandel 2008; De Roo et al. 2008). After long-term memory is established by the initial stages of consolidation, later rounds of consolidation may be needed to extend the persistence of the memory (Wittenberg and Tsien 2002; Bekinschtein et al. 2007). Consolidated memories may also be modified when they are retrieved (Dudai 2006; Alberini and Taubenfeld 2008). Retrieving a memory by exposure to the conditioned stimulus, or by re-experiencing aspects of the original training, can destabilize it and initiate an additional protein-synthesis-dependent process of memory stabilization (Nader 2003; Dudai 2006; Alberini and Taubenfeld 2008). The aim of this communication is to explore whether an experience that alone does not cause long-term memory can make a memory more persistent and can also destabilize a consolidated memory and makes it labile.Training Aplysia californica with inedible food until they stop responding to the food initiates long-term memory that can be measured as a reduction in the time to stop responding. Long-term memory is present 1, 2, and 7 d after a single training session (Schwarz et al. 1991). We examined some of the parameters of training that lead to persistence of memory by determining whether shorter training sessions also produce a persistent long-term memory.As in previous studies (Schwarz et al. 1991; Botzer et al. 1998; Lyons et al. 2005), Aplysia californica were trained with inedible food, the seaweed Ulva wrapped in plastic net. This food induced biting, leading to food entering the mouth. Animals then attempted to swallow the food. The netted food cannot be swallowed and it became lodged in the buccal cavity, producing repetitive failed swallowing responses. Food eventually left the buccal cavity. The experimenter continued to hold the food against the lips, inducing further biting responses, entries into the mouth, and failed swallows. As training proceeded many bites failed to cause entry of food into the mouth. When food did enter the mouth it stayed within for progressively shorter periods, eliciting fewer attempted swallows. In all animals, food was in the mouth eliciting failed attempts to swallow for at least 100 sec of the initial training, since previous experience showed that such animals almost always show long-term memory. Animals in which food was not in the mouth for 100 sec during training were discarded. A full training session until animals stop responding to food requires 10–25 min of training (Fig. 1). Such a training session caused long-term memory measured after 24 h or after 48 h (Fig. 1A). Memory was measured by comparing the time to stop responding to inedible food during training to the time to stop responding when animals were tested 24 or 48 h later, in a procedure identical to that during training. All tests of memory were performed using a blind procedure in which the experimenter was unaware of the previous training procedure.Open in a separate windowFigure 1.Memory 24 and 48 h after different training procedures. The figure shows the time to stop responding in a group of naïve animals trained with a full training session (training continued until animals stop responding to food), as well as the time to stop responding on memory trials 24 and 48 h later (N = 38 naïve animals [naïve animals were run as controls for the various other groups and data from the naïve animals were then combined. There was no significant difference between the various groups of naïve animals: P = 0.405, F_(5,32) = 1.053]); (A) N = 13 animals tested 24 h after a full training; N = 6 animals tested 48 h after a full training; (B) N = 9 animals tested 24 h after a 5-min training; N = 17 animals tested 48 h after a 5-min training; (C) N = 14 animals tested 24 h after a 3-min training; (D) N = 7 animals tested 48 h after a 5-min training, plus an additional 3-min training 24 h later. A one-way analysis of variance showed significant differences between the seven groups shown (P < 0.001, F_(6,97) = 11.95). A post-hoc test (Student-Newman-Keuls, α = 0.05) showed that there were no significant differences between the time to stop in naïve animals and in animals tested 48 h after a 5-min training, or in animals tested 24 h after a 3-min training, indicating that these treatments did not cause memory. By contrast, the times to stop in these three groups were significantly different from that in the other four groups, which were not significantly different from one another. These findings indicate significant memory 24 or 48 h after a full training, as well as 24 h after 5 min of training, and 48 h after 5 min of training with 3 min of reminder training, with no differences in memory after these 4 treatments. Standard errors are shown.We examined whether abbreviated training also caused long-term memory. First, we examined long-term memory when training is stopped after 5 min. During the first 5 min of a full training session, food is in the mouth, and animals are attempting to swallow for a mean of 70% of the time in the mouth during a full training session. Attempts to swallow are an integral part of the training (Katzoff et al. 2006). We confirmed an earlier finding (Botzer et al. 1998) that a 5-min training produced long-term memory measured 24 h after the training (Fig. 1B). However, we have now found that training animals for only 5 min did not cause long-term memory measured 48 h after training (Fig. 1B). A training session that was stopped after 3 min did not give rise to long-term memory measured at 24 h (Fig. 1C), indicating that the last 2 min of a 5-min training are necessary for the production of 24 h memory. These findings allowed us to examine the possible effects of an experience which itself does not cause memory, 3 min of training on memory persistence and memory lability.A 5-min training session gives rise to memory after 24 h, but not after 48 h, whereas additional training gives rise to 48 h memory. Must the additional training take the form of continuing to train animals during the initial training session, or can an additional 3 min of training that itself does not cause 24 h memory enhance the effect of a 5 min of training? To test the ability of a 3-min training to enhance memory, animals were trained with either a full training session, or with a training session that was abbreviated after 5 min. One group of animals trained for 5 min received an additional 3-min training 24 h after the initial training, whereas another group did not. Memory was tested 24 h later, 48 h after the initial training. Animals receiving a full training, and animals receiving a 5-min training plus a reminder consisting of a 3-min additional training, displayed 48 h memory (Fig. 1D; for a fuller presentation of this experiment, see Supplemental material), whereas animals receiving only a 5-min training, with no additional training, showed no 48 h memory. These data show that a 3-min training, which is itself ineffective in producing memory 24 h later, can lead to significant memory when it follows a 5-min training, which itself would not produce 48 h memory.In other learning tasks it has been shown that long-term memory is not a unitary process. An earlier round of protein synthesis is necessary for 24 h memory, but a more persistent memory (>24 h), and the synaptic plasticity underlying it, are dependent on later rounds of protein synthesis (Giustetto et al. 2003; Bekinschtein et al. 2007; Miniaci et al. 2008). We therefore examined whether 24 and 48 h memory differ in their dependence on protein synthesis adjacent to training, or on protein synthesis 6 h after training. The protein-synthesis inhibitor anisomycin was injected into the hemolymph either 10 min before training or 6 h after training. Animals were injected with a 1 cc solution of anisomycin at a concentration that caused a 10 µM concentration within the animals. This concentration blocks protein synthesis in ganglia (Schwartz et al. 1971). Controls were injected with 1 cc of artificial seawater (ASW–NaCl 460 mM, KCl 10 mM, CaCl₂ 11 mM, MgCl₂ 55 mM, and NaHCO₃ 5 mM). Treatment with anisomycin just before training blocked 24 h memory, but anisomycin treatment 6 h after training did not prevent the appearance of 24 h memory, indicating that 24 h memory is consolidated within the first 6 h after training (note that Fig. 2A shows the percent change in time to stop responding during the test of memory, with respect to the time to stop during training). By contrast, 48 h memory was blocked by anisomycin treatment 6 h after training (Fig. 2B), whereas treatment with ASW did not block 48 h memory.Open in a separate windowFigure 2.Protein synthesis dependence of 24 and 48 h memory. Data shows the memory test 24 or 48 h after training as a percent change [−([train-test]/train) × 100] from the initial training session. All tests of memory were after a full training session until animals stopped responding to the food. Memory was tested via two-tailed paired t-tests, in which the time to stop responding in a given animal during the initial training was compared with the time to stop responding 24 or 48 h later, when animals were tested. A significant decrease in the time to stop responding was used as an indicator of memory. Treatments showing such a decrease are marked (*). Standard errors are shown. (A) Anisomycin treatment blocks 24 h memory when applied just before training, but not when applied 6 h after training. Control animals (N = 8) not treated with anisomycin displayed significant 24 h memory (P < 0.001, t₍₇₎ = 10.15). Anisomycin applied 10 min before the training (N = 9) blocked memory, as shown by a lack of significant decrease in the time to stop between the initial training and the test after 24 h (P = 0.96, t₍₈₎ = 0.05). By contrast, application of anisomycin 6 h after the initial training (N = 7) did not block memory measured 24 h after training, as shown by significant savings after 24 h (P = 0.05, t₍₆₎ = 2.45). (B) Forty-eight hour memory is dependent on a later stage of protein synthesis. Anisomycin (N = 26), but not ASW (N = 5) applied 6 h after training blocks 48 h memory, as shown by significant memory after treatment with ASW (P = 0.007, t₍₄₎ = 5.08) but not with anisomycin (P = 0.49, t₍₂₅₎ = 0.69). However, a 3-min reminder training 24 h after the training (N = 12) rescues 48 h memory after it was blocked by anisomycin treatment 6 h after training (P = 0.01, t₍₁₁₎ = 2.93).Memory 48 h after training is dependent on protein synthesis 6 h after training. As shown above, 3 min of training can establish 48 h memory after a 5-min training that alone is too brief to establish 48 h memory. Can 3 min of training also establish 48 h memory after it has been blocked by inhibiting protein synthesis at 6 h? To test this possibility, animals were trained with inedible food until they stopped responding, and were then treated with either ASW or anisomycin 6 h later. One group of animals also received 3 min of training with inedible food 24 h after the initial training. When tested 48 h after training, memory was present in animals treated with ASW and in animals that had received the additional 3-min training, but not in animals receiving only the anisomycin treatment (Fig. 2B). This finding indicates that a 3-min training that is effective in causing 48 h memory after 5 min of training can also rescue 48 h memory after it has been blocked by anisomycin.If 3 min of training saves a blocked 48 h memory, it could also amplify an already-formed 48 h memory. We tested the effect on 48 h memory of a 3-min training 24 h after a full training session, which alone produces 48 h memory. There was no significant difference in memory between animals tested 24 h after training, and animals receiving a 3-min retraining, and then tested 48 h after training, indicating that the 3-min training did not affect the already consolidated 48 h memory (Fig. 3, cf. column 1 and column 3).Open in a separate windowFigure 3.Brief training makes memory labile. Twenty-four hours after training with inedible food, animals were treated with anisomycin, or with ASW. The animals treated with ASW (N = 9), as well as one of the two groups treated with anisomycin (N = 9), were also given a 3-min training session 10 min later. Memory was then tested again 24 h later, 48 h after the training, by comparing the time to stop measured 48 h after training with the time to stop measured during the original training session, using a two-tailed paired t-test. A significant decrease in the time to stop responding was used as an indicator of memory. Data are also shown for 24 h memory after a full training session, to allow a test of whether a 3-min retraining at 24 h improves memory tested at 48 h. Treatment with anisomycin alone (N = 8) did not block 48 h memory (P = 0.002, t₍₇₎ = 4.66). The 3-min reminder training plus treatment with ASW also did not block 48 h memory (P = 0.001, t₍₈₎ = 5.02). By contrast, when the 3-min reminder training was preceded by anisomycin treatment, there was no significant difference between training and the 48 h test (P = 0.27, t₍₈₎ = 1.18), indicating that the 3-min reminder allowed the anisomycin to block 48 h memory. Note that there is also no significant difference in savings after the 3-min training following ASW treatment and 24 h memory following training (P = 0.13, t₍₁₀₎ = 1.63), indicating that the 3-min training on day 2 alone did not affect memory.The 3-min training does not affect an already-formed 48 h memory, but does cause effects 24 h later if the memory is not fully consolidated. Formation of a long-term memory requires protein synthesis, and recalling a consolidated memory can make a memory sensitive to inhibitors of protein synthesis (Nader et al. 2000). Does a 3-min training 24 h after an initial full training initiate a renewed dependence of 48 h memory on protein synthesis? Animals were trained to criterion and then subjected to either anisomycin, or ASW, 24 h later. Both groups then received a 3-min reminder training 10 min later. Memory was then assessed 24 h later (48 h after initial training). There was significant memory 48 h after the training after treatment with ASW, but not after treatment with anisomycin, indicating that the 3-min reminder training restored the ability of anisomycin to block memory (Fig. 3).Our data have shown that 3 min of training with inedible food alone does not produce long-term memory (Fig. 1), and does not amplify an already established 48 h memory (Fig. 3). Nonetheless, 3 min of training induces a protein-synthesis-dependent state in which memory can be modified (Figs. 2, ​,3).3). After a previous training leading to only 24 h memory (Fig. 1), or after a treatment blocking 48 h memory, the 3-min training makes the memory more persistent (Fig. 2). In addition, after a 48 h memory has already been consolidated, 3 min of training makes the memory labile, so that it can be blocked by inhibitors of protein synthesis (Fig. 3).Reconsolidation is a process by which a consolidated memory returns to a protein-synthesis-dependent labile state by retrieving the memory (e.g., Nader et al. 2000; Sangha et al. 2003; Kemenes et al. 2006). In learning that food is inedible 3 min of training makes the memory labile. There has been much speculation on the possible function of memory reconsolidation. It has been suggested that the destabilization of memory by retrieval could function as a means to update and modify the magnitude and the persistence of the original memory trace (Dudai 2002, 2006). There is good evidence that reconsolidation can update (Rodriguez-Ortiz et al. 2005; Morris et al. 2006), strengthen (Frenkel et al. 2005; Tronson et al. 2006; Lee 2008), or modify (Rossato et al. 2006) memories, as well as block them (Nader et al. 2000), but the possible role of reconsolidation in regulating memory persistence has not been examined. Indeed, many of the previous studies on reconsolidation used a conditioned stimulus (CS) alone as a reminder, and thereby caused extinction along with reconsolidation (Eisenberg et al. 2003; Pedreira and Maldonado 2003), rather than causing a strengthening of memory. Previous studies have indicated that after memory is initially consolidated, its persistence is dependent on successive waves of protein synthesis (Giustetto et al. 2003; Bekinschtein et al. 2007; Miniaci et al. 2008) and that reactivation of a memory improves its persistence (Spear 1973). Later waves of protein synthesis affecting persistent memories could be modulated or modified by retrieving a memory.We have taken advantage of a memory task affecting Aplysia feeding to examine the possibility that memory retrieval via a brief additional training affects later waves of protein synthesis, and thereby affects the persistence of a memory. We found that pairing a brief additional training with block of protein synthesis blocks memory (Fig. 3). We also found that a more persistent memory measured at 48 h is dependent on a longer training session (Fig. 1), and a later wave of protein synthesis (Fig. 2) than is 24 h memory. Retrieval of memory by a 3-min retraining can establish 48 h memory even when the latter portion of training is absent (Fig. 1), or when the later wave of protein synthesis is blocked (Fig. 2). This finding suggests that the initial experience required to establish 48 h memory, as well as the wave of protein synthesis elicited by this experience, can be deferred. A later experience, and a later wave of protein synthesis, can substitute for the absence of experience in the initial training, or for the blocked wave of protein synthesis. In addition to creating a more persistent memory, a 3-min retraining also makes a consolidated memory sensitive to blocking by inhibitors of protein synthesis (Fig. 3). Thus, a function of experiences that permit reconsolidation may also be to extend the persistence of a memory.In our study we cannot exclude the possibility that the 3-min retraining does more than just retrieve memory, and thereby make it labile. The increased persistence of memory could be caused by processes initiated by the 3-min training, such as increased consolidation, that are not related to the ability of the added training to make the memory labile. Indeed, previous studies have suggested that memory consolidation and reconsolidation may utilize different molecular mechanisms (Taubenfeld et al. 2001; Lee et al. 2004; Alberini 2005). The different molecular mechanisms activated by consolidation or reconsolidation have been used to classify the process by which retrieval affects memory, although use of the molecular markers to classify a behavioral process is controversial (Nader et al. 2005). Such studies have shown that changing a memory when it is retrieved sometimes utilizes a molecular mechanism for consolidation (Tronel et al. 2005), and sometimes for reconsolidation (Lee 2008). In the learning task utilized we have not examined separate molecular markers for consolidation and reconsolidation, raising the possibility that a 3-min retraining affects persistence by activating molecular mechanisms that are specifically associated with a separate round of consolidation, rather than activating reconsolidation.Memory reconsolidation also affects a number of other molluscan learning paradigms (Sekiguchi et al. 1997; Sangha et al. 2003; Gainutdinova et al. 2005; Kemenes et al. 2006). In some, the memory becomes labile when animals are exposed to a CS alone, which does not itself create memory, whereas in others the memory becomes labile only when the CS is paired with an unconditioned stimulus (US), as in the original training (see Eisenhardt and Stollhoff 2008). In our experiments, memory was made labile by a second training session, which was shorter than that needed to itself create memory. It will be of interest to determine whether components of the experience alone in which animals learn that food is inedible do not give rise to memory, such as lip stimulation alone (Schwarz et al. 1988), can also give rise to reconsolidation, although they would not cause a more persistent memory.Where in the Aplysia nervous system are molecular changes related to reconsolidation and the persistence of memory localized? The earliest molecular changes leading to long-term memory that a food is inedible (e.g., increased expression of C/EBP and sensorin), are localized to the buccal ganglia, specifically to a population of mechanoafferents (Levitan et al. 2008a,b). It is possible that the later molecular changes leading to a persistent memory are also localized to these neurons, similar to the localization to the same sensory neurons of earlier and later molecular events leading to long-term facilitation underlying sensitization in Aplysia (Miniaci et al. 2008). However, it is also possible that the later phases of consolidation, and reconsolidation, may arise by molecular changes localized to other sites with the Aplysia nervous system, similar to the movement of earlier and later phases on long-term declarative memories from the hippocampus to the neocortex (Squire and Kandel 1999). Future studies will need to examine these points.