Place-preference task in a 2D VR environment.

(A) 2D VR setup. (B) Bird’s-eye view of the virtual environment. Various landmarks surrounded a circular arena, and a fixed start location (‘St’) was at the center. Reward zones are illustrated with white dashed lines for visualization purposes. (C) Place-preference task paradigm. A trial started with one of six pseudorandomly chosen start directions (‘Trial Start’). In this example, the rat started the trial facing the northeast (NE) direction, highlighted in green. Subsequent navigation is illustrated here with the associated scene (‘Navigation’). A dot on the gray trajectory indicates the rat’s current location and the black arrow describes the head direction. When the rat arrived at a reward zone, honey water was delivered within 8 seconds, with the visual scene frozen (‘Reward’). Finally, a gray screen appeared, denoting an inter-trial interval; if the rat remained still (<5 cm/s) for 5 seconds, the subsequent trial began (‘ITI’).

Common body-turning behavior of rats after learning.

(A) The reference frame of the virtual environment. The six start directions are illustrated with the red high-value zone (180°) and blue low-value zone (0°). On the right, the departure circle (DC) is denoted with a purple dashed line, and the start direction is marked with a black arrowhead and a green arrow. (B) Overall changes in scene direction over the normalized distance between the start location and the DC (left). Each colored line indicates the median change of scene direction in trials with each start location, and red and blue arrowheads mark high- and low-value zone centers, respectively. The 0°-to-360° range was repeated in the ordinate of the plot to capture rotational movements in opposite directions (positive and negative directions for clockwise and counterclockwise rotations, respectively). The gray lines on the right show the rat’s trajectory within the DC. These examples were excerpted from the first and last days of pre-training of a single rat. (C) Individual examples of scene directions and trajectories in the novice session. Scene direction change for each direction is drawn separately (top) for individual trials. The black arrowhead indicates that specific start direction. Trajectories within the DC (middle) and the whole arena (bottom) are also illustrated according to the indicated color code. Mean travel distance and latency are shown below the VR arena trajectory. (D) Same as (C), but for the expert session.

Learning index for efficient navigation during pre-surgical training.

(A) Changes in departing direction (DD) with learning. (i) Schematic of DD (purple dot), with the departure circle shown as a dashed line. (ii) Distribution of DDs in pre- and post-learning sessions from all rats (rose plots). Gray denotes the pre-learning session, whereas purple indicates the post-learning session. Mean vectors are illustrated as arrows with the same color scheme, and their lengths are indicated at the upper right side of the plot. (iii) Schematic of the DD-deviation angle (angle between the high-value zone center and the DD) and comparisons of DD-deviation angles between pre- and post-learning sessions. Each dot represents data from one rat. (B) Same as (A), but for perimeter-crossing direction (PCD; green dot). The perimeter is drawn as a green dashed circle. **p < 0.01.

Cannula implantation locations and schedules for training and drug injection.

(A) Cannula positions marked. (i) Example of bilaterally implanted cannula tracks in Nissl-stained sections in the dHP and iHP. (ii) Tip locations illustrated in the atlas, with different colors for individual rats (n = 8). (B) Training schedule. Rats were divided into two groups (n = 4/group) to counterbalance the injection order for the main task and probe test.

Changes in navigational pattern with each drug condition.

(A) Sample trajectories in each drug condition. Black arrowheads indicate the start direction and the gray line shows the trajectory for each trial. (B) Mean high-value zone visit percentage for each drug condition. Gray, green, and orange each indicate PBS, dMUS, and iMUS sessions, respectively. (C) Average running speed. (D) Number of perimeter crossings. For the PBS session, dPBS and iPBS sessions were first tested for significant differences between sessions; if they were not different, they were averaged to one PBS session for analysis purposes. *p < 0.05.

dHP and iHP inactivation differentially affect efficient goal-directed navigation.

(A) Grouped comparison of DD in each drug condition. (i) Distributions of DDs in each drug condition (rose plots) and a comparison of their mean directions. Gray plots, PBS sessions; green plots, dHP inactivation; orange plots, iHP inactivation. Red and blue arcs indicate high- and low-value zones, respectively. Statistically significant differences in mean vectors, illustrated as arrows, are indicated with asterisks. The mean directions of all four conditions were first compared together (Watson-Williams test); a post hoc pairwise comparison was subsequently applied if the average mean vector length of the two sessions was greater than 0.45. The number on the upper right side of the plot shows the length of the mean vector. (B, C) Changes in mean vector length (B) and deviation angles from the high-value zone center (C) of the DD in each drug session. *p < 0.05, **p < 0.01, ***p < 0.001.

Accuracy of goal-directed navigation is more severely impaired with iHP inactivation.

(A-C) Same as Figure 6, except showing PCD. *p < 0.05, **p < 0.01, ***p < 0.001.

Goal-directedness and navigational capacity are unaffected by drug infusion.

(A) Object-guided navigation task as a probe test. (i) A flickering object appeared on either the left (‘Left trial’) or right (‘Right trial’) side of the screen. The start location is marked with a yellow dot, with a white arrow indicating the start direction, which remained the same for both trial types. (ii) Example of trajectories in one session. Blue and red lines represent trajectories from left and right trials that directly arrived at the reward zones, whereas gray lines indicate failed trials. Green dashed lines denote reward zones. (B) Comparison of the proportion of each drug condition’s direct hit trials (both left and right).