Table 1.
Fig 1.
An electrode capacitance is calculated from the actual reading of the electrode at t and the base-line value.
The values from the 12 electrodes are then used to calculate the x, y position of the mouse centroid (left panel). A PA bout is initiated when x, y changes in successive samples and continuous until the mouse is still again (no change in x, y between successive samples). The time-series of x, y coordinates during a movement bout is used to plot the bout trajectory (right panel) and to decide if it is a MOTS or locomotor bout. Inserted into the track diagram (right panel) is how the cage floor was divided into a front(FF)-rear(RF) area (green line and text) and a central(CF)-peripheral(PF) area of equal size (red line and text), respectively. For further information see text.
Table 2.
Fig 2.
A Boxplots showing number of PA and rest bouts per day for single housed female mice. Rest is further divided into short and long rest, and PA in local movements (MOTS) and locomotor bouts. B Boxplots showing time spent in rest, long rest, short rest, and PA, in MOTS, and locomotion as fraction of total time. Values indicated are mean, 1st and 3rd quartiles and range. Mean value has been indicated with a blue circle.
Fig 3.
A Cumulative plot of fraction of the total number of bouts (ordinate) vs bout duration (s, abscissa, logarithmic scale) for all bout types (black line with SD as grey shaded area), rest bouts (black line with blue shaded area indicating SD), MOTS (black line over green area indicating SD), and locomotion bouts (black line on red shaded area indicating SD). B Cumulative percentage of total time (ordinate) plotted vs. bout duration (s, abscissa, logarithmic scale). Coloured area and code for bout type are the same as in A. Dotted line in A and B indicates number of bouts (˜90% in A) having a duration <10 s and their combined fraction of total time (<25% in B). A and B, data from single housed female mice.
Table 3.
Fig 4.
A and B show impact by day post cage change (dp 0–13; abscissa) and cage change cycle (CC 1–3; colour coded grey, red, and green) of fraction of total time spent in bouts of PA (A) and (B) long rest (ordinates). The relative effect size of dp and CC is shown in S5 Fig in S1 File. In A (model: PA ˜dp * CC), the major impact is by dp (p = 6.2E-11) with no contribution by CC (p = 0.16). B shows that CC has no significant impact on time in long rest (p = 0.54) while dp has a strong effect (p = 4.3E-11) (model: long rest˜dp*CC). A and B, data from single housed female mice.
Fig 5.
Pattern of long-rest episodes during lights on (ZT 0–12) and lights off (ZT 12–24) (DL 12:12) for the mice S1, S7 and S10 housed in isolation.
Long rest bouts indicated by blue colour. Bouts disrupting long-rest periods in black/green. Ordinate is 12h lights on to the left and 12h lights off to the right. Columns are day post cage change (dp) where 0 is the day of the cage change (CC; red arrow indicates the time for CC) for lights on (to the left) and lights off (to the right). Dp1 the day after CC and dp13 the day before the next CC. See S6 Fig in S1 File for corresponding data of the other 7 single housed female mice.
Table 4.
Fig 6.
(A) shows cumulative distance per 12 hrs during lights off (black line) and lights on (blue line), respectively. Values are average across the 10 animals during ZT 0–12 and ZT 12–24 each day with standard error indicated by bars. (B) shows corresponding data for average speed (±SEM) during lights on (blue) and lights off (black). A and B, data from single housed female mice.
Fig 7.
A-C show longest recorded locomotor bout for mouse S1, S9 and S10, respectively, during the observation period. In D-F, the corresponding speedograms are depicted. Track plots and speedograms by the trajar package in R. For tracks and speedogram of the other 7 single housed female mice see S8 Fig in S1 File.
Fig 8.
Densitograms showing the distribution of bout starting coordinates during 24h on dp0 and dp13 (columns) in cage change cycles 1 (A) and 3 (B) for the S7 mouse. Cage front and rear and left (L) and right (R) have been indicated. Top row of panels show MOTS (green) and locomotion (red) bouts, while lower row of panels show the corresponding data for short (orange) and long (blue) rest bouts. Please see S1 File for corresponding metrics of the other female mice kept in isolation.
Fig 9.
Reduction (%) of bouts dp13 vs dp0.
Fig 10.
A Boxplots showing the preference for bout initiation in the frontal field of the cage floor on dp0 and dp13 through cage change cycles 1–3. In B and C, the relative frequency of long rest and locomotor bouts, respectively, starting in the frontal field of the cage floor have been indicated. D-F show the corresponding boxplots when the cage floor was divided into a central and peripheral field of equal size (see also Fig 1). A -C show data from single housed female mice.
Fig 11.
A shows fraction of daily distance (m) per hours during the LD cycle for each single housed female mouse (thin coloured lines) and the average across the ten cages (thick black line). B shows the EAD per hour when the animals are active during the LD cycle. Individual mice indicated by thin coloured lines, average across the group is the thick black line. C show the average fraction of each hour the mice spend in long rest (blue), short rest (orange), MOTS (grey) and in locomotion (yellow) across the LD cycle. Abscissa is Zeitgeber time with lights on 0–12 and lights off 12–24, the shift on to off is marked by interrupted vertical lines.
Fig 12.
A-B Boxplots of fraction of total file spent at rest (no electrode activation) (A) and long rest (B) for the cohorts of female and male mice housed at different density (Fx1, Mx2, Fx3 etc). C-D show boxplots of duration and density of long rest bouts assessed by the EAD metric. As in A-B, female and male cohorts housed at different densities have been indicated. Statistics by ANOVA with model rest/long rest ˜ density*sex; significance of each factor has been indicated in the plots.
Fig 13.
A-B Cumulative plots of time in PA assessed by EAD and the number of electrode activations observed (abscissa) as fraction of file time (ordinate) for each cohort of female (A) and male (B) cages (starting point of curve represent all bouts of rest (R). Housing density (x1, x2, x3, and x4) has been colour coded (key in panel A-B). Solid line represents cohort average value across cages and weeks of recording. The shaded area with the same colour indicates the standard deviation. Interrupted vertical lines indicate cut point values for rest (R) and change in electrode activations s-1from one, two, four and eight or more electrodes activated.
Table 5.
Fig 14.
Panels show the average fraction of each hour spent in PA (yellow) and at rest (blue) across the LD cycle for male and female mice housed at different densities.
Rest i.e., no electrode activation and PA when electrode activations occur. Abscissa is Zeitgeber time with lights on 0–12 and lights off 12–24, the shift on to off is marked by interrupted vertical lines.
Fig 15.
A show fraction of file time spent in PA (≥ 1 electrode activated s-1) across cages per day (dp) of the cages change cycles 1–3 for male and female mice housed at different densities (x1: n = 1, x2 n = 2, x3 n = 3 and x4 n = 4). Comparison of female (red) and male (blue) mice at density n = 3 and n = 4, respectively, revealed a significant difference at density n = 4 but not when density = 3 (model: Time in PA ˜ sex * dp * CC; density = 4 F = 36.6; n = 3 F = 0.07). See also Fig 10 in S1 File for plot of relative effect size of dp and CC for males and females at densities n = 3 and n = 4, respectively. B show fraction of file time spent in long rest bouts (<1 electrodes activated s-1 and≥40 s duration of bout) across cages and cages-change cycles (CC 1–3) per day (dp0-dp13) for male (blue) and female (red) mice housed at different densities (x1 to x4). Comparison of female and male mice at density n = 3 and n = 4, respectively, revealed a significant difference in synchronized long rest bout time at density n = 4 but not when n = 3 (model: Time in long rest ˜ sex * dp * CC; n = 4 F = 7.6; n = 3 F = 0.79). For plot of relative effect size of dp and CC at these densities see Fig 10B in S1 File. C show average number of unique electrodes activated s-1 across cages per day of the cages change cycles 1–3 for male (blue) and female (red) mice housed at different densities (x1 to x4). Comparison of female and male mice (model: No unique electrodes ˜ sex * dp * CC) revealed a significant difference at density = 4 (F = 41, p = 1.9E-5) but not at x3 (F = 1; p = 0.33). For plot of relative effect size of dp and CC at these densities see Fig 10C in S1 File.
Fig 16.
The panels left-to-right show relative effect size (RTE, ordinate) on average daily EAD by housing density (n), days post cage change (abscissa; dp0-13) and cage-change cycle (CC 1–3; colour coded black, red, and green, in each panel) within sexes (group of panels). Whole model: EAD ˜ N * dp * CC; for females F = 163.7, p = 7.6E-15; for males F = 28, p = 3.3E-7. In both sexes, density (n) had the strongest impact on EAD, followed by dp, while impact by cage change cycle was only significant in females. NB: In males only two cycles could be compared across densities. Note that at N = x4 EAD show a biphasic trajectory across all cycles in both sexes (red arrows, for further information see text).
Table 6.