C57BL/6J is one of most commonly used inbred strains of mice, especially as a genetic background in mutant mouse lines 
. Confirming a few previous studies that have reported PIT in C57BL/6J and (mixed or congenic) C57BL/6J-background mutant mice using comparable procedures as employed here [e.g.]
, [ 33]
, we found robust PIT in this strain. This was evidenced by increased instrumental response rates during presentations of an auditory cue previously associated with reward delivery, compared to presentations of a cue not associated with the reward. Furthermore, this cue-induced instrumental responding was specifically directed to the lever that had delivered reward during training although, interestingly, there was an increase in responding on the control lever during the no-CS periods; potentially reflecting a type of explorative response driven by the ‘release-from-stimulus-control’ during periods when no cue was available to predict either the availability or absence of reward. It is worth noting that, in addition to showing increased lever pressing during the CS+ (PIT), C57BL/6J also showed robust CS+-controlled Pavlovian approach or goal-tracking responses (head entries). The expression of PIT (instrumental responding) and Pavlovian approach have been reported to compete at the behavioral level 
. Additionally, as PIT and CS-controlled goal-tracking rely on separate neurobiological mechanisms 
, observing both in the same mouse and during the same single test allows for powerful dissociations based on strain and/or genetic mutation [e.g.]
, [ 35,36]
. Collectively, our PIT data demonstrate strong formation of associative representations underlying incentive motivation and robust expression of stimulus-controlled reward-seeking behaviors in the C57BL/6J strain.
This conclusion was bolstered by the performance of this strain in our instrumental stimulus-response paradigm where C57BL/6J mice readily learned to correctly respond to a visual stimulus on a touchscreen to obtain a food reward. In turn, when reinforcement was omitted, instrumental responding efficiently extinguished, indicating that a S-R association had been established during training. We have reported similar patterns in C57BL/6J and 129/SvImJ inbred mice 
and C57BL/6J-background mutants 
. A novel observation here was that (extinguished) responding was robustly reinstated. Reinstatement was produced either by brief presentation of the US and the tone/light compound cue associated with the reward (conditioned reinforcer, CR), by brief US presentation followed by presentation of the CRs on all remaining trials, or by presentation of the CRs alone on all trials. Interestingly, the magnitude of responding during these different reinstatement conditions was not different in the C57BL/6J strain, indicating that exposure to the CRs alone was an effective re-energizer of responding, but that the continued presentation of the CRs after an initial exposure to the reward itself was not sufficient to further augment responding. The lack of any differences or additive effects could arguably reflect a behavioral ceiling effect, although this seems less likely as responding remained at around thirty percent of the maximum. Additional, controlled experimentation would be required to explore this issue further. It would also be useful in future experiments to include a CS− condition in order to confirm that CS-induced reinstatement of responding was explicitly due to its associative history with reward.
An interesting and somewhat surprising finding was that C57BL/6J mice were insensitive to outcome devaluation caused by repeated pairing of the food US with sickness induced by LiCl injection. This was indicated by the fact that neither instrumental responding (lever pressing) nor discriminated approach or goal-tracking behavior (magazine head-entries) were reduced in mice having undergone LiCl-food paired devaluation, relative to non-devalued control mice. It is important to emphasize, however, that the absence of any instrumental devaluation effect was not simply an artifact of a failure of mice to form an association between the food US and the experience of malaise, because mice in the devalued group clearly showed a persistent aversion to consuming the freely available reward in the home cage and in the operant chambers themselves. By definition then, these findings suggest that the instrumental reward-seeking response is, at least in part, habitual and driven by processes that are separate and divorced from the outcome.
On the other hand, the absence of a devaluation effect contrasts with previous reports of clear outcome devaluation induced using sensory-specific satiety, rather than LiCl pairings, in C57BL/6J-background mice [e.g.]
, [ 30]
. In these studies, instrumental lever press responses were markedly reduced in mice that had been sated with the reward (but not a different natural reinforcer) prior to the probe test. Certain instrumental training variables, including the schedule of reinforcement 
and the overall number of reinforcers earned 
, are known to determine sensitivity to outcome devaluation. However, these factors cannot account for the discrepancy between previous studies because we controlled for reinforcers earned and used the same training procedure and random ratio schedule previously shown to be effective in producing satiety-induced devaluation in C57BL/6J-background mice 
. Whatever the case, these contrasting findings indicate that differences exist between sensitivity to malaise- and satiety-induced devaluation that render the former ineffective in reducing instrumental lever press behavior in C57BL/6J mice.
This conclusion was not limited to C57BL/6J mice, as we found that DBA/2J mice also failed to show altered responding following reinforcer devaluation (again, despite clear evidence of the formation of a successful food-sickness pairing). To our knowledge, this is the first published report of a reinforcer devaluation procedure used with this mouse strain. DBA/2J mice have been widely used in behavioral neuroscience and, within addiction research, have been heavily studied because of an aversion (relative to C57BL/6J) to orally consumed alcohol 
. This strain (along with C57BL/6J) also has an important role in behavior genetic studies as one of the parental strains of the BXD recombinant panels employed to identify sources of genetic variation underlying, for example, behavioral responses to abused drugs including alcohol 
, nicotine 
and psychostimulants 
In addition to insensitivity to malaise-induced reward devaluation, we found that DBA/2J mice were similar to C57BL/6J mice in that they by and large showed robust PIT, extinction and reinstatement. The purpose of the current study was not to cross-compare strains as strains were not tested in a fully counterbalanced design, precluding direct statistical comparison. However, testing was done under identical conditions and informal visual comparison of the data suggests that DBA/2J mice were similar to C57BL/6J mice on the majority of behavioral measures. One exception was that DBA/2J mice seemed slower to extinguish the instrumental response – requiring more sessions to extinguish this response than to acquire it (C57BL/6J mice showed the opposite pattern). The DBA/2J strain also failed to show significant reinstatement of this response when exposed to conditioned reinforcers alone. While this suggests weak conditioned reinforcement in this paradigm, it cannot simply be explained by a more general failure to form cue-reward relationships, per se
, as PIT was clearly intact in these mice. Additionally, although both rely broadly on intact corticostriatal circuitry function, numerous studies in rats and mice show that PIT and conditioned reinforcement are dissociable at the systems, receptor and molecular levels 
. Thus, further studies will be needed to clarify whether the DBA/2J strain has a genuine deficit in certain forms of reinstatement.
The third strain we characterized on these tasks was BALB/cJ. BALB/cJ has been well-studied for its heightened anxiety-like behavior and stress reactivity in comparison to, for example, C57BL/6J [e.g.]
, [ 53]
. As such, this strain could prove valuable to studies aimed at identifying genetic and molecular factors modulating stress effects on cognitive and executive deficits in addiction [for further discussion see 56]. We found that BALB/cJ mice displayed good PIT, extinction and reinstatement. During the PIT probe, BALB/cJ mice did not engage in exploration of the inactive lever during the CS− and no-CS periods, which is reminiscent of their reduced tendency to explore in anxiety-provoking environments. The BALB/cJ strain also exhibited poor discriminated Pavlovian approach behavior during the PIT probe, i.e., these mice did not engage in significantly more reward-seeking during the CS+ than the CS−.
As with the other two strains, instrumental lever pressing was undiminished by reinforcer devaluation. Interestingly, however, there was a significant decrease in magazine entries in devalued BALB/cJ mice (although the strength of the LiCl-induced illness-food pairing seemed relatively weak in these mice and was not expressed on the long-term retention test). This indicates that at least one component of the behavioral repertoire of BALB/cJ mice in this test was sensitive to the current reward value. But, again, instrumental lever pressing and Pavlovian-approach behaviors are dissociable processes 
, and their differential sensitivity to outcome devaluation is not unprecedented. For example, Nelson and Killcross found that repeated amphetamine treatment rendered rats insensitive to either malaise- or satiety-induced devaluation of lever pressing but not head entry behavior 
. As discussed by these authors, magazine entry behavior may be more resistant to changes in reward value either due to its proximity to the reward and/or because it has a Pavlovian approach component [see 60]
. In this context, our data suggest that the BALB/cJ strain may provide an interesting genetic model to explore the nature of this dissociation in future studies.
In summary, the current study describes a set of paradigms for assaying various operant-based reward-related behaviors in three of the most commonly used inbred mouse strains. We describe a procedure for demonstrating PIT, and a method for studying acquisition, extinction and multiple forms of reinstatement of an instrumental touchscreen response. While we were unable to demonstrate malaise-induced devaluation of an instrumental response in any of our strains (although Pavlovian approach responses were sensitive to diminished outcome value in BALB/cJ mice) we found no reason to conclude that mice were unable to form the necessary food-malaise association and the negative results more likely point to more complex factors needing additional study. These procedures provide a useful platform for future studies using the mouse as a model species to elucidate the critical neural, molecular and genetic factors subserving reward-related behaviors, and ultimately provide new insights into maladaptive manifestations of motivated behaviors such as drug addiction.