presents the individual microdialysis probe placements for all rats. In general, the probes were successfully implanted in the targeted brain area, either the accumbens core (n = 6), accumbens shell (n = 7), DLS (n = 7), or DMS (n = 8).
Schematic representation of microdialysis probe placements for rats in Groups Core (n = 6; black), Shell (n = 7; gray), DLS (n = 7; white), and DMS (n = 8; horizontal stripes). Numbers represent distance (mm) of coronal section from bregma.
Acquisition of lever pressing
As can be seen in , rats from the four probe placement conditions acquired comparable rates of instrumental performance. An ANOVA using surgical group (core, shell, DMS, and DLS) and session (1–7) as factors found a significant main effect of session (F6,144 = 99.72; p < 0.001), but found no effect of surgical group (F3,24 = 1.16; p = 0.35) or session by group interaction (F18,144 = 0.97; p = 0.50).
Acquisition of lever pressing. Mean (± SEM) lever presses performed per minute for rats in Groups Core, Shell, DLS and DMS.
Baseline dialysate DA levels
Mean DA levels during pre-testing baseline periods were 0.83 nM (SEM ± 0.15) for the core, 0.73 nM (SEM ± 0.13) for the shell, 1.46 nM (SEM ± 0.17) for the DLS, and 0.96 nM (SEM ± 0.14) for the DMS. Analysis of these data revealed a significant main effect of probe placement (F3,26 = 4.72; p = 0.01). Bonferroni post-hoc tests found that basal DA levels were significantly higher in the DLS than in core (p = 0.05) or shell (p < 0.01). No other differences reached significance, indicating that DA levels in the DMS were intermediate between those observed in the DLS and nucleus accumbens.
Conditioning-induced DA efflux in the nucleus accumbens
Tests 1 and 3 allowed us to measure changes in extracellular DA concentrations as rats lever pressed for food reward in a state of high food motivation. Although task performance was comparable across these two tests, a mixed ANOVA using probe placement (core and shell) and test (1 and 3) as factors found that rats with accumbens probe placements responded at a marginally higher rate in Test 3 than in Test 1 (F1,11 = 3.41; p 0.092), which may have resulted from the additional conditioning given during the test sessions. No effect of placement (F1,11 = 0.24; p = 0.634) or placement by test interaction (F1,11 = 0.20; p = 0.667) was detected. Analysis of the number of rewards earned during these tests found no effect of test (F1,11 = 0.24; p = 0.632) or placement (F1,11 = 0.06; p = 0.808), and found no test by probe interaction (F1,11 = 0.11; p = 0.742).
Instrumental conditioning resulted in an elevation of accumbal DA during the session, which returned to near baseline levels after the session ended (see ). These data were analyzed using a mixed ANOVA with probe placement, test, and time (23 × 3-min samples) serving as factors. The ANOVA detected a significant effect of time (F22,242 = 11.37; p < 0.001), confirming that DA levels changed over the course of the session. There was no effect of test (F1,11 = 0.94; p = 0.354) or probe placement (F1,11 = 0.26; p = 0.619). Nor was there a significant test by group (F1,11 = 0.01; p = 0.929), test by time (F22,242 = 0.83; p = 0.682), or test by time by group interaction (F22,242 = 0.75; p = 0.785).
Figure 3 DA efflux in the core (left panel) and shell (right panel) of the nucleus accumbens of hungry rats lever pressing for food pellets. Black lines represent mean (± SEM) DA concentrations (expressed as percentage of baseline; i.e., average for bins (more ...)
Although the magnitude of conditioning-induced DA efflux in the accumbens was similar for the two hungry tests when the data were averaged across animals, our multi-test experimental design allowed us to explore the relationship between conditioning-induced DA efflux and important experimental variables using an approach that minimizes between-subjects variability (e.g., probe function). Specifically, we examined if individual differences in the average DA response during instrumental conditioning (average of bins 1–10 for each subject during test sessions) between hungry tests were correlated with differences in either the number of lever presses performed or the number of pellets earned between the two sessions. Furthermore, since reinforcement was delivered in a probabilistic manner according to a RR-10 schedule, rats experienced variability in the number of responses needed to obtain reward (i.e., total number of presses performed per session/total number of rewards obtained per session) between the two tests, allowing us to investigate the relationship between DA and the effort (or response cost) required to gain each outcome. For each variable, Test 1 values were subtracted from Test 3 values to produce a difference score. presents the mean (SEM) and range of difference scores for each of these variables. We found that changes in DA efflux in the nucleus accumbens (n = 13) between Tests 1 and 3 were not significantly correlated (Pearsons, two-tailed) with changes in response rate (r = −0.049; p = 0.874) or changes in the number of rewards earned (r = 0.432; p = 0.140) between the two tests. Therefore, our data do not support the view that accumbal DA mediates a simple response-activating influence over instrumental performance, or else DA levels should have varied with response vigor. Nor does DA appear to simply track the number of actions performed or number of pellets earned. However, changes in DA were found to be negatively correlated with changes in the number of presses required to obtain reward (r = −0.580; p = 0.038; see ), such that the DA response during conditioning dropped as the work requirement increased, and vice versa. This correlation was significant when the data from rats with accumbens core placements were separately analyzed (n = 6; r = −0.851; p = 0.032), indicating that the modulation of conditioning-induced DA efflux by response requirement in this structure is particularly strong. Although no correlation was found for rats with shell placements (n = 7; r = −0.567; p = 0.184), this lack of effect should be interpreted with caution, particularly given the significant correlation found when both probe placement groups were included in the analysis.
Table 1 The mean difference in conditioning-induced DA efflux (average of bins 1–10), total lever presses performed, total rewards earned, and ratio of responses per reward (i.e., average response requirement) between the two Hungry Tests (Test 3 – (more ...)
Figure 4 Scatter plot showing the relationship between changes in conditioning-induced DA efflux and response requirement (i.e., the mean number of lever presses required to earn each reward) across hungry tests (Test 3 – Test 1) for rats in Groups Shell (more ...)
To assess the effects of general satiety on tonic DA signaling and instrumental performance, we contrasted data collected during tests conducted when the rats were hungry (average of Tests 1 and 3) with the data from Test 2, which was conducted after the rats had been sated for 2h on home chow. As shown in , this satiety induction procedure was generally effective in reducing the rate of lever press performance (main effect of motivational state: F1,11 = 25.79; p < 0.001). No effect of probe placement (F1,11 = 0.65; p = 0.438) or state by placement interaction (F1,11 = 0.93; p = 0.355) was detected. An ANOVA performed on the number of rewards earned during hungry and sated tests found that rats earned significantly fewer rewards when sated than when hungry (F1,11 = 28.22; p < 0.001). No effect of placement (F1,11 = 0.41; p = 0.536) or state by placement interaction (F1,11 = 0.99; p = 0.340) was detected.
Figure 5 Effect of satiety on conditioning-induced DA efflux in the core (left panel) and shell (right panel) of the nucleus accumbens. Black lines represent mean (± SEM) DA concentrations (expressed as percentage of baseline; i.e., average for bins −3, (more ...)
In general, the effects of instrumental conditioning on accumbal DA efflux also appeared to be sensitive to downshift in motivational state (see ). A mixed ANOVA performed on these data detected a marginally significant main effect of motivational state (F1,11 = 4.24; p = 0.064), a significant effect of time (F22,242 = 12.92; p < 0.001) and, more importantly, a significant state by time interaction (F22,242 = 1.78; p = 0.020), indicating that the time course of conditioning-induced DA efflux was affected by the rats’ motivational state at test, with rats exhibiting an attenuated DA response when they were sated. There was no main effect of probe placement (F1,11 = 0.14; p = 0.718), nor was there any interaction between placement and state (F1,11 = 0.04; p = 0.843), placement and time (F22,242 = 0.62; p = 0.910), or between these three variables (F22,242 = 0.98; p = 0.49).
The correlational analysis described above indicated that accumbal DA does not exert a direct response-activating effect on instrumental performance, or track the number of responses performed or rewards earned. However, if accumbal DA is involved in the motivational control of instrumental performance, one might expect the effects of satiety on DA efflux to vary with the behavioral effects of satiety. To assess this possibility, we examined whether individual changes in the average DA response during instrumental conditioning (average of bins 1–10 for each subject) between Sated (Test 2) and Hungry tests (data averaged across Tests 1 and 3) correlated with differences in the rate of responding or number of pellets earned between these conditions. For each variable, the mean value for the Hungry tests was subtracted from the value for the Sated test to generate a difference score; therefore, a negative difference score indicates that the Sated value was lower than the Hungry value. The mean difference scores (SEM) and the range scores are presented in . When the data were collapsed across sites, a marginally significant correlation was detected between changes in accumbal DA efflux and changes in response rate (r = 0.514; p = 0.072). Although a similar relationship was apparent between changes in DA and changes in the number of rewards earned, this correlation also failed reach the conventional threshold for significance (r = 0.455; p = 0.118). Further analysis limited to accumbens shell placements, however, found significant positive correlations for both response rate (r = 0.827; p = 0.022; see ) and rewards earned (r = 0.778; p = 0.039; see ), which were highly correlated with one another (r = 0.973; p < 0. 001) since rewards were delivered according to a ratio schedule. Changes in core DA efflux were not correlated with either variable (response rate: r = −0.241; p = 0.646, rewards earned: r = −0.243; p = 0.643). Thus it appears that, at least for the accumbens shell, the suppressive effects of satiety on conditioning-induced mesolimbic DA efflux vary with its behavioral effects.
Table 2 The mean difference in conditioning-induced DA efflux (average of bins 1–10), total lever presses performed, and total rewards earned between the Sated and Hungry Tests (Test 2 – average of Tests 1 and 3). SEMs are presented in parentheses. (more ...)
Figure 6 Scatter plot showing the relationship between the effect of satiety (Sated Test – Hungry Test) on conditioning-induced DA efflux and its effects on lever press performance (Panel A) and on the number of rewards earned (Panel B) for rats in Groups (more ...)
Conditioning-induced DA efflux in the dorsal striatum
A mixed ANOVA using probe placement (DLS, DMS) and test (1 and 3) as factors found that rats with dorsal striatal probe placements responded at similar rates across the two hungry tests (F1,13 = 0.87; p = 0.368). No effect of placement (F1,13 = 0.20; p = 0.662) or placement by test interaction was detected (F1,13 = 0.26; p = 0.621). Analysis of the number of rewards earned in Tests 1 and 3 found no effect of test (F1,13 = 1.03; p = 0.328) or placement (F1,13 = 0.16; p = 0.692), and found no interaction between these factors (F1,13 = 1.16; p = 0.300). Although somewhat more modest than in the nucleus accumbens, dorsal striatal DA levels also increased as hungry rats lever pressed for food reward during Tests 1 and 3 (see ), an effect that was observed in both lateral and medial regions. A mixed ANOVA using probe placement group, test, and time as factors detected a significant main effect of time (F22,286 = 2.83; p < 0.001), confirming that DA levels increased during these test sessions. There was no effect of test (F1,13 = 0.07; p = 0.791) or probe placement (F1,13 = 0.48; p = 0.502), nor was there any significant interactions between group and test (F1,13 = 0.02; p = 0.905), group and time (F22,286 = 1.16; p = 0.286), or between the combination of these three variables (F22,286 = 0.81; p = 0.720).
Figure 7 DA efflux in the DLS (left panel) and DMS (right panel) of hungry rats lever pressing for food pellets. Black lines represent mean (± SEM) DA concentrations (expressed as percentage of baseline; i.e., average for bins −3, −2, and (more ...)
Although dorsal striatal DA levels significantly increased during tests conducted when rats were hungry, this effect did not appear to be associated with any of the behavioral variables being recorded. Changes in the DA response (n = 15, collapsed across placements) to instrumental conditioning between Tests 1 and 3 were not correlated with changes in response rate (r = 0.077; p = 0.785), number of rewards earned (r = 0.112; p = 0.691), or response requirement (r = 0.220; p = 0.431).
shows the effects of satiety on lever pressing and conditioning-induced DA efflux in the dorsal striatum. Rats with dorsal striatal placements also responded at a significantly lower rate when sated than when hungry (F1,13 = 8.43; p = 0. 012). There was no effect of probe placement (F1,13 = 0.87; p = 0.369) and no state by placement interaction (F1,13 = 0.73; p = 0.407). Analysis of the number of rewards earned in hungry and sated tests found a main effect of motivational state (F1,13 = 10.02; p = 0.007). No effect of placement (F1,13 = 0.26; p = 0.622) or interaction between state and placement (F1,13 = 0.06; p = 0.816) was detected. Although DA levels in the dorsal striatum appeared to increase slightly at the beginning of the sated test session, these changes were much less persistent than those observed when the rats were tested hungry (average of Tests 1 and 3). This interpretation was supported by the statistical analysis, which although failing to detect a significant overall effect of motivational state (F1,13 = 1.89; p = 0.192), did find a significant state by time interaction (F22,286 = 1.62; p = 0.041). There was no effect of group (F1,13 = 0.89; p = 0.362), nor was there any significant interactions between group and state (F1,13 = 0.001; p = 0.975), group and time (F22,286 = 1.25; p = 0.206), or between group, state and time (F22,286 = 0.51; p = 0.969). Although it appears that conditioning-related DA efflux in the dorsal striatum was modulated by motivational state, unlike in the accumbens, subject-specific changes in DA efflux between Hungry and Sated tests were not significantly correlated (n = 15; collapsed across placements) with changes in response rate (r = −0.108; p = 0.702) or rewards earned (r = 0.011; p = 0.969).
Figure 8 Effect of satiety on conditioning-induced DA efflux in the DLS (left panel) and DMS (right panel) of the nucleus accumbens. Black lines represent mean (± SEM) DA concentrations (expressed as percentage of baseline; i.e., average for bins −3, (more ...)