Group characteristics at baseline
There were no significant differences between the sham, anodal, cathodal groups on age, Body Mass Index (BMI) and time of testing, F(2,42) < 1.43, p > .250. Furthermore, groups were matched on gender (χ2(2) = .27, p = .874), as well as use of hormonal contraception and cycle phase in the female sample (Fisher’s Exact tests: p > .373). Groups did not differ significantly at baseline on (log)cortisol (F(2,42) = 1.68, p = .199), TMD (F(2,42) = .05, p = .950) and VAS (H(2) < 3.22, p > .200). Finally, there were no group differences on most trait measures (F(2,24) < 2.50, p > .094). Scores obtained on the openness (BFI-44), appraisal of emotions and social skills (SSREIS) subscales were significantly different between groups (F(2,42) > 3.26; p < .048). However, the variance in the total cortisol output (AUCg) could not be significantly explained by group membership (dummy coded factor) (R2 = .049, F(2,42) = 1.09, p = .347), or by either of the three personality measures when these were additionally entered (together) in the regression model (R2 = .123, F(5,39) = 1.09, p = .380). We concluded that these baseline differences were unlikely to affect performance on the saccadic adaptation task, as they did not affect participants’ endocrine output. Therefore, differences in adaptation metrics were expected to arise from the experimental manipulation (Table 1).
Cortisol levels and mood
Logarithmic transformation was applied to normalize the cortisol data (Fig. 3). A 2 × 2 ANOVA with Group factor (Sham, Cathodal, Anodal) and Time (baseline, t + 1, t + 10, t + 30) as the within-subjects factor revealed a main effect of time, F(1,55) = 24.84, p < .001, η2p = .372. There was no main effect of group (F(1,42) = 1.04, p = .361, η2p = .047) and no interaction (F(3,55) = .36, p = .757, η2p = .017). Cortisol levels decreased from the beginning of the experiment to the final sample (t(44) = 6.36, p < .001). There were no differences in the total cortisol output (AUCg) amongst the 3 groups, F(2,42) = 1.09, p = .347.
We also assessed changes in mood. A 2 × 2 ANOVA with Group factor (Sham, Cathodal, Anodal) and Time (TMD pre-tDCS, TMD post-tDCS) as the within-subjects factor also demonstrated a main effect of time, F(1,42) = 14.69, p < .001, η2p = .259. There was no group effect (F(1,42) = .07, p = .934, η2p = .003) and no significant interaction (F(2,42) = 1.77, p = .182, η2p = .078). Follow-up comparisons showed an overall improvement in mood after tDCS (M = 9.69, SD = 19.30), compared to baseline (M = 18.33, SD = 21.99), t(44) = 3.78, p < .001. There were no significant changes in mood on all VAS scales across groups (Wilcoxon ranked tests: Z > -1.34, p > .180). Within the cathodal group, participants felt less tense–pressured (M = 1.28, SD = .61) post-tDCS compared to baseline (M = 1.93, SD = 1.07), Z = -2.46, p = .014. All other within group comparisons were not significant (p > .084).
In summary, tDCS polarity did not affect cortisol levels or subjective mood. There was an overall improvement in mood and a decrease in cortisol output post-tDCS.
Saccadic baseline performance
We evaluated whether stimulation polarity influenced saccade metrics at baseline. Saccadic gain, duration, velocity and latency were independently submitted to three-way ANOVAs with Block (Pre1, Pre2), Direction (leftward, rightward), as the within-subjects factors, and Group (Sham, Cathodal, Anodal) as the between-subjects factor.
For gain and velocity, analyses revealed main effects of direction (gain: F(1,42) = 17.80, p < .001, η2p = .298; velocity: F(1,42) = 62.11, p < .001, η2p = .597). Rightward saccades had higher gains across groups and averaged blocks, t(44) = 4.29, p < .001 (Fig. 4a), and higher velocity across averaged blocks in each stimulation group: Sham (t(15) = 4.31, p = .001); Cathodal (t(13) = 4.81, p < .001); Anodal (t(14) = 4.86, p < .001). Analysis on velocity also yielded a group effect with greater overall velocity in Sham (F(2,42) = 5.31, p = .009, η2p = .202). Given that the velocity group difference was present when no stimulation was applied (no significant group x block interaction, p = .825), we interpret this finding as a pre-existing difference that is independent of stimulation polarity (Fig. 4c). Saccadic duration was also not affected by tDCS polarity and there were no baseline differences (F < 3.19, p > .082) (Fig. 4b). Finally, for latency, we found a significant block x group interaction (F(2,42) = 4.95, p = .012, η2p = .191). However, follow-up comparisons between groups over averaged directions revealed non-significant differences at Pre1 (p > .190) or Pre2 (p > .545) among the three groups (Fig. 4d).
Given the absence of stimulation polarity effects on baseline adaptation metrics, change values in the adaptation and post-adaptation sequences were calculated relative to the mean preadaptation values (Pre1 and Pre2).
Effects of tDCS stimulation polarity on adaptation time-course and aftereffects
Adaptation rates were first evaluated by fitting a linear slope to the gain change values of 120 adaptation trials for each participant. No significant differences were found between the adaptation slopes in the sham (M = .05, SD = .08), cathodal (M = .005, SD = .08) and anodal (M = .07, SD = .08) groups (F(2,42) = 2.50, p = .094). However, mean values were indicative of milder adaptation slopes in the cathodal group. This was further investigated over 10 time points (bins).
A two-way ANOVA with Group factor (Sham, Cathodal, Anodal) and Time measured over 10 levels (adaptation bins) showed a progressive increase in saccade size in all groups (time effect: F(4,168) = 5.19, p = .001, η2p = .110). Adaptation rates were also significantly different between groups (group effect: F(2,42) = 3.64, p = .035, η2p = .148). There was no time x group interaction (F(8,168) = 1.52, p = .152, η2p = .068). Bonferroni-corrected pairwise comparisons were employed to explore group differences throughout the adaptation sequence. Anodal participants had greater gains compared to the Sham group at bins 3 (t(29) = − 2.53, p = .046) and 4 (t(29) = − 2.50, p = .039). Compared to the Cathodal group, the Anodal group also exhibited higher gain changes at bins: 7 (t(27) = 2.62, p = .036); 9 (t(27) = 2.79, p = .023); 10 (t(27) = 2.93, p = .016). All other comparisons were not significant (p > .068) (Fig. 5).
Finally, the postadaptation phase was implemented to evaluate aftereffects in the absence of saccadic error. In a two-way ANOVA evaluating the effects Group (Sham, Cathodal, Anodal) and Time (Post 1, Post 2), gain change was not significantly different between the two blocks (time effect: F(1,42) = 1.12, p = .296, η2p = .026). Across blocks, we found a significant group effect (F(2,42) = 3.32, p = .046, η2p = .137). Group differences were independent of time (interaction: F(2,42) = .50, p = .611, η2p = .023). As there was no time effect, the two Post blocks were averaged. Bonferroni-corrected comparisons on averaged blocks revealed that aftereffects were significantly greater in the Anodal group compared to the Cathodal group (t(27) = 2.58, p = .041). There were no significant differences between the active stimulation groups and participants undergoing sham stimulation (p > .517).
To summarize, results suggest that stimulation polarity differentiated between the two active groups in the second half of the adaptation sequence. Compared to the Cathodal group, were changes were small from bin1 to bin10 (1.6%), participants in the Anodal group showed on average an 8.4% increase in gain. This difference was also present in Post. However, increased gain change in the Anodal group was also present early in the adaptation sequence, suggesting higher overall values. Furthermore, saccadic gain under active stimulation was not significantly different from the control group.