Spatially restricted inhibition of cholinergic interneurons in the dorsolateral striatum encourages behavioral exploration

Abstract

When pursuing desirable outcomes, one must make the decision between exploring possible actions to obtain those outcomes and exploiting known strategies to maximize efficiency. The dorsolateral striatum (DLS) has been extensively studied with respect to how actions can develop into habits and has also been implicated as an area involved in governing exploitative behavior. Surprisingly, prior work has shown that DLS cholinergic interneurons (ChIs) are not involved in the canonical habit formation function ascribed to the DLS but are instead modulators of behavioral flexibility after initial learning. To further probe this, we evaluated the role of DLS ChIs in behavioral exploration during a brief instrumental training experiment. Through designer receptors exclusively activated by designer drugs (DREADDs) in ChAT-Cre rats, ChIs in the DLS were inhibited during specific phases of the experiment: instrumental training, free-reward delivery, at both times, or never. Without ChI activity during instrumental training, animals biased their responding toward an "optimal" strategy while continuing to work efficiently. This effect was observed again when contingencies were removed as animals with ChIs offline during that phase, regardless of ChI inhibition previously, decreased responding more than animals with ChIs intact. These findings build upon a growing body of literature implicating ChIs in the striatum as gate-keepers of behavioral flexibility and exploration.

Keywords: DREADDs; basal ganglia; habit; learning; reward.

FIGURE 1

Histological results. (a) Schematic representation of DIO-hM4D(Gi)-mCherry expression in the dorsolateral striatum (DLS) of rats from Group Vehicle and Group CNO (n = 40). (b) Schematic representation of DIO-mCherry expression in the DLS of rats from Group CNO control (n = 10). Numbers indicate distance from bregma in mm. Coronal slices adapted from Paxinos and Watson (2009). (c) Representative image showing DIO-hM4D(Gi)-mCherry in the DLS. (d) Group assignments by experiment phase, with CNO administration noted. During FR1 training, injections were not given. Upon FR3 training, animals were split into three groups (Vehicle, CNO, and CNO control). Group Vehicle and Group CNO received hM4D(Gi) DREADDs tagged with mCherry while Group CNO control received infusions mCherry alone during surgery. At RT60, groups were split once again, to control for CNO-exposure history. CNO, clozapine N-oxide

FIGURE 2

Behavioral training results. (a) Presses per reinforcer. All three groups of rats, Group CNO, Group Vehicle, and Group CNO control, pressed the active lever equivalently as measured by the number of presses made per reinforcer over FR3 training days. (b) Presses per minute. The average Group CNO press rate as a measure of presses made per minute increased compared to the two control groups over FR3 training days. (c) Magazine entries per reward. The overall average entries made per reward decreased over training days. (d–f) Bout-type probabilities by training day. Three types of bouts are presented, one-bout (d), two-bout (e), and three-bout (f) in white, orange, and purple, respectively. Shapes indicate group assignment (diamond = CNO control, square = Vehicle, circle = CNO). There is a drop in the average one-bout probability, and a concomitant rise in two- and three-bout probabilities, in Group CNO but not in control groups. FR1 bout probabilities, before animals were exposed to CNO, are presented to the left of the dashed line to show stability in tendencies between FR1 training and the 1st day of FR3 training. Error bars represent ± SEM. (g) Bout frequencies by group assignment on FR3 Day 3. Group Vehicle and Group CNO show similar one-press (white), two-press (orange), three-press (purple), and four-press and greater (gray) bout frequency tendencies on the final day of fixed-ratio training. Group CNO used the one-press bout strategy less than control animals while increasing the use of the other bout strategies

FIGURE 3

Behavioral testing results. (a) Schematic representing how animals with hM4D(Gi) were split into groups from FR3 training (two groups) to RT60 testing (4 groups). Splitting animals as such allowed to control for cholinergic interneuron inhibition history when assessing behavior in test. CNO control animals received a virus that lacked the DNA for the hM4D(Gi) DREADD but were given CNO injections during each phase of the experiment. (b) RT60 presses per reward (PPR). The average press rate of each group as a measure of PPR over all RT60 test sessions. (c) Percent change in PPR behavior among groups. Press behavior on Day 2 of RT60 testing was compared to the press behavior of Day 1 and is presented as a percent change. Animals with cholinergic interneurons off-line at the time of test (left) decreased lever press behavior more than controls, regardless of CNO-exposure history. This effect was not present if animals were grouped by ChI inhibition during training (right). Bars are colored to indicate which groups were pooled together for analysis. (d) Percent change in magazine entries made per minute across all groups. Similar to the results presented in c, magazine entry rates on Day 2 of test were compared to magazine entry rates on Day 1 and are presented as a percent change. There are no observed effects of group on magazine entry behavior, indicating that entry behavior remained steady across these two test sessions. (e) One-press bout probability changes between the final training day and the first test day by RT60 group assignment. (f) Three-press bout probability changes between the final training day and the first test day by RT60 group assignment. (g) Changes in one-press bout probabilities during the second RT60 test session as a percentage of the first RT60 test session, by group assignment. (h) Changes in three-press bout probabilities during the second RT60 test session as a percentage of the first RT60 test session, by group assignment. All error bars represent ± SEM