Wireless Optogenetic Stimulation of Oxytocin Neurons in a Semi-natural Setup Dynamically Elevates Both Pro-social and Agonistic Behaviors

May combine two claims.

The wireless optogenetic device is head-mounted, magnetically activated, compact, simple to construct, and suitable for use in group-living mice in an enriched semi-natural arena over several days.

“Here, we present a head-mounted, magnetically activated device for wireless optogenetic manipulation that is compact, simple to construct, and suitable for use in group-living mice in an enriched semi-natural arena over several days.”

May combine two claims.

Repeated optogenetic activation of PVN-OT neurons in groups of male mice produced dynamic behavioral effects: an initial phase of increased social interest, interpreted as pro-social behavior, followed by a concomitant increase in agonistic behavior.

“Combining the use of wireless optogenetic devices with an enriched social setting, we found dynamic behavioral effects of repeated optogenetic activation of PVN-OT neurons in groups of male mice. Following an initial phase of increased social interest, which can be interpreted as pro-social behavior, we observed a concomitant increase in agonistic behavior.”

• AssumptionRests on an assumption: The semi-natural arena and group-housed conditions provide an ethologically relevant social context for studying OT effects.

May combine two claims.

The device comprises a fiber optic cannula, an LED powered by two "multi-drain" watch batteries, and a magnetic field-dependent reed-switch.

“comprises a fiber optic cannula, an LED powered by two “multi-drain” watch batteries, and a magnetic field-dependent reed-switch”

Probably not a substantial claim: may not pass S3 (load-bearing).

All parts of the device are commercially available.

“All parts of the device are commercially available”

May combine two claims.

The wireless device is small (<1 cm3, approximately 1.5 g with dental cement coating) and head mountable.

“It is small (<1 cm3, approximately 1.5 g with dental cement coating) and head mountable”

May combine two claims.

A 1 s blue light pulse (488 nm) induced membrane depolarization in PVN-OT neurons, sometimes accompanied by action potential firing, which slowly decayed over approximately 30 min to resting membrane potential.

“In each recorded cell, the application of a 1 s blue light pulse (488 nm) induced membrane depolarization, accompanied in some cases by action potential firing. This depolarization slowly decayed with time (approximately 30 min) to the resting membrane potential”

May combine two claims.

The mean amplitude of SSFO-induced depolarization was 10.1 ± 0.66 mV, and deactivation with a 10 s green light pulse (561 nm) completely reversed this effect back to baseline levels (488 nm: 10.1 ± 0.66 mV versus 561 nm: −0.4 ± 0.49 mV; p = 6.1 × 10−5, n = 14 cells).

“The mean amplitude of SSFO-induced depolarization was 10.1 ± 0.66 mV. Deactivation of the SSFO with a 10 s green light pulse (561 nm) completely reversed this effect back to baseline levels (Figure 2D; 488 nm: 10.1 ± 0.66 mV versus 561 nm: −0.4 ± 0.49 mV, relative to pre-stimulation resting membrane potential; one-tailed exact paired permutation test, p = 6.1 × 10−5, n = 14 cells).”

May combine two claims.

In the 5 brains assessed, an average of 75.5% ± 12% of tdTomato+ cells in the PVN co-expressed OT and 96% ± 4% of OT+ cells co-expressed tdTomato.

“In the 5 brains assessed, an average of 75.5% ± 12% of tdTomato+ cells in the PVN co-expressed OT and 96% ± 4% of OT+ cells co-expressed tdTomato”

May combine two claims.

The device produces sufficient light intensity for effective activation of considerable target volumes using SSFO, with an estimated 1.75 mm3 of brain tissue illuminated above 8 microwatt×mm-2, the lowest reported functional light intensity for SSFO.

“This provided an estimated 1.75 mm3 of brain tissue being illuminated with light intensity above 8 microwatt×mm–2, the lowest reported functional light intensity for SSFO”

• cites Yizhar et al., 2011 sourceNot checked yet

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May combine two claims.

SSFO-mediated photoactivation of PVN-OT neurons led to a significant increase in plasma OT in Cre+ but not Cre- mice following a single light pulse and following two consecutive days of repeated activations.

“In both cases, SSFO-mediated photoactivation of PVN-OT neurons led to a significant increase in plasma OT in Cre+ but not Cre– mice (Figures 2I–2J; single activation: Cre+, two-tailed paired permutation test, p = 0.0156, n = 7; Cre–, two-tailed paired permutation test, p = 0.7891, n = 9. Repeated activation: Cre+, two-tailed paired permutation test, p = 0.00778, n = 8; Cre–, two-tailed paired perm”

• AssumptionRests on an assumption: Plasma OT levels reflect central OT release from PVN-OT neurons.

May combine two claims.

There were no significant differences in locomotion (total distance covered or percentage of distance covered while engaging in social/non-social behavior) between groups.

“we quantified locomotion and found no significant differences in either the total distance covered by the mice or the percentage of distance covered while engaging in social/non-social behavior”

May combine two claims.

Photoactivation significantly increased the length of self-grooming bouts and shortened the latency to the first bout in Cre+ but not Cre- mice on both activation days.

“Photoactivation significantly increased the length of self-grooming bouts, and shortened the latency to the first bout, in Cre+ but not Cre– mice on both activation days”

May combine two claims.

The mean duration of grooming bouts was highest in Cre+ mice treated with saline; Kruskal-Wallis revealed a significant effect of treatment (p = 0.0302), with post hoc differences between Cre+|saline and the two other groups approaching significance (p = 0.06315 and p = 0.06254).

“The mean duration of grooming bouts was highest in Cre+ mice treated with saline (Figure 3C). Kruskal-Wallis analysis revealed a significant effect of treatment on this measure (chi-square test = 7, df = 2, p = 0.0302); post hoc Dunn’s test found that the differences between the Cre+|saline group and the two other groups approached significance (Cre+|saline versus Cre– p = 0.06315; Cre+|saline”

May combine two claims.

Time spent alone outside the nest decreased in Cre+ mice but increased in Cre- mice from baseline to activation day 1 (p = 0.0028).

“Similarly, time spent alone outside the nest decreased in Cre+ mice but increased in Cre– from baseline to activation day 1 (Figure 4E; two-tailed permutation test, p = 0.0028).”

May combine two claims.

On the first activation day, PC1 scores were positively correlated with affiliative behaviors such as rates of approach and contact between conspecifics, and negatively correlated with time spent alone outside the nest.

“on the first activation day, PC1 scores were positively correlated with affiliative behaviors, such as rates of approach and contact between conspecifics, and negatively correlated with time spent alone outside the nest, a prototypical asocial behavior”

May combine two claims.

Approach and chase behavior, as well as time alone outside the nest, were independent of dominance rank.

“Importantly, approach and chase behavior, as well as time alone outside the nest, were independent of dominance rank (Figures S4C–S4G; Pearson correlation).”

May combine two claims.

The first 2 principal components cumulatively explained 41% (activation day 1 versus baseline) and 44% (activation day 2 versus baseline) of the total variance, significantly higher than expected by chance (PC1 p = 4.2 × 10−7, PC2 p = 0.00088).

“The first 2 principal components (PC1 and PC2) cumulatively explained 41% (activation day 1 versus baseline) and 44% (activation day 2 versus baseline) of the total variance in the data, which was in both cases significantly higher than expected by chance, as calculated by randomly permuting the data (see STAR Methods, Quantification and Statistical Analysis; Figures S3F and S3G; two-tailed perm”

May combine two claims.

On the second day of manipulation, the increase in social interest is accompanied by an increase in agonistic behaviors such as anogenital sniffing and aggressive chases.

“On the second day of manipulation, however, this trend is accompanied by an increase in agonistic behaviors such as anogenital sniffing and aggressive chases.”

Probably not a substantial claim: may not pass S3 (load-bearing).

The resting membrane potential of SSFO-expressing neurons was 60.2 ± 1.5 mV.

“The resting membrane potential (Vm) of SSFO-expressing neurons was 60.2 ± 1.5 mV.”

Classified as probably not a substantial claim.

The wireless optogenetic device can be easily adapted for use in a variety of behavioral paradigms that are normally hindered by tethered light delivery or a limited environment.

“Our wireless optogenetic device can be easily adapted for use in a variety of behavioral paradigms, which are normally hindered by tethered light delivery or a limited environment.”

Probably not a substantial claim: may not pass S4 (based).

Semi-natural setups for measuring behavior are incompatible with traditional optogenetic approaches, which use tethered light delivery systems.

“However, semi-natural setups for measuring behavior are incompatible with traditional optogenetic approaches, which use tethered light delivery systems.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

OT has long been known to be involved in social behaviors and is recently being explored as a therapeutic candidate for autism spectrum disorders (ASDs).

“OT has long been known to be involved in social behaviors and is recently being explored as a therapeutic candidate for autism spectrum disorders (ASDs)”

• cites Jurek and Neumann, 2018 sourceNot checked yet

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• cites Ooi et al., 2017Not checked yet

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Classified as probably not a substantial claim.

The social salience hypothesis suggests that OT acts to regulate the salience of external social cues, rather than strictly promoting affiliative behaviors.

“These findings gave rise to the social salience hypothesis, which suggests that OT acts to regulate the salience of external social cues, rather than strictly promoting affiliative behaviors”

• cites Shamay-Tsoory and Abu-Akel, 2016 sourceNot checked yet

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• cites Steinman et al., 2019 sourceNot checked yet

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May combine two claims.

SSFO upon activation by a single light pulse leads to prolonged (approximately 30 min) sub-threshold depolarization, sensitizing the cell to afferent inputs rather than inducing firing directly.

“we chose to use the Stabilized Step Function Opsin (SSFO), which upon activation by a single light pulse leads to prolonged (approximately 30 min) sub-threshold depolarization (Yizhar et al., 2011), sensitizing the cell to afferent inputs rather than inducing firing directly.”

• cites Yizhar et al., 2011 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

PVN OT neurons are the source of most OT forebrain projections.

“We therefore targeted the OT neurons in the PVN, which are the source of most OT forebrain projections”

• cites Knobloch et al., 2012 sourceNot checked yet

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May combine two claims.

Several studies have demonstrated a reduction in aggressive behaviors upon treatment with exogenous OT or an increase in such behaviors in animals genetically lacking OT or OTR.

“Several studies have employed the RI paradigm to demonstrate a reduction in aggressive behaviors upon treatment with exogenous OT (Calcagnoli et al., 2013, Calcagnoli et al., 2014, Calcagnoli et al., 2015, Jong et al., 2014, Karpova et al., 2016) or an increase in such behaviors in animals genetically lacking OT or OTR (Dhakar et al., 2012, Pagani et al., 2015, Ragnauth et al., 2005, Takayanagi et”

• cites Calcagnoli et al., 2013 sourceNot checked yet

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• cites Dhakar et al., 2012 sourceNot checked yet

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• cites Ragnauth et al., 2005 sourceNot checked yet

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May combine two claims.

Even a mild depolarization results in systemic OT release and the OT content of the neurons is not depleted following 2 days of activation.

“This indicates that even a mild depolarization results in systemic OT release and, conversely, that the OT content of the neurons is not depleted following 2 days of activation.”

• AssumptionRests on an assumption: Plasma OT levels reflect central OT release from PVN-OT neurons.

Classified as probably not a substantial claim.

These findings have led to the notion that OT generally reduces aggression.

“These findings have led to the notion that OT generally reduces aggression.”

Probably not a substantial claim: may not pass S4 (based). May combine two claims.

The potential confounds of receptor desensitization during pharmacological treatment, and developmental effects in genetic KOs, can be eliminated with an optogenetic approach.

“Importantly, the potential confounds of receptor desensitization during pharmacological treatment, and developmental effects in genetic KOs, can be eliminated with an optogenetic approach.”

May combine two claims.

In the context of a group of familiar males, prolonged PVN-OT neuronal activation leads to a dynamic behavioral shift, comprising an immediate elevation in social interest (interpretable as pro-social behavior) and a delayed increase in agonistic behavior.

“Taken together, these results suggest that in the context of a group of familiar males, prolonged PVN-OT neuronal activation leads to a dynamic behavioral shift, comprising an immediate elevation in social interest, which can be interpreted as pro-social behavior, along with a delayed increase in agonistic behavior.”

Classified as probably not a substantial claim. May combine two claims.

Observations of mice colonies in a visible burrow system have shown that approach toward the back of a conspecific more often induces flight than approach to the front and thus may reflect an offensive motivation.

“Observations of mice colonies in a visible burrow system have shown that approach toward the back of a conspecific more often induces flight than approach to the front and thus may reflect an offensive motivation”

• cites Arakawa et al., 2007 sourceNot checked yet

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May combine two claims.

Understanding of the ethology of mice and rats in the wild is still lacking, and this gap has been acknowledged as a major challenge in the field.

“Unfortunately, understanding of the ethology of mice and rats in the wild is still lacking, and this gap in knowledge and methodology has been acknowledged as a major challenge in the field (Anderson and Perona, 2014, Blanchard et al., 2013, Kondrakiewicz et al., 2019).”

• cites Anderson and Perona, 2014Not checked yet

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• cites Blanchard et al., 2013 sourceNot checked yet

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• cites Kondrakiewicz et al., 2019 sourceNot checked yet

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Probably not a substantial claim: may not pass S2 (checkable). May combine two claims.

Reductionist behavioral tests, though more controllable, raise questions regarding translatability; if a task is far enough removed from an animal's natural environment or behavioral repertoire, it may reflect neural or behavioral responses not used naturally.

“Though the widely used reductionist tests have the advantage of better controllability, they still raise questions regarding translatability—if a behavioral task is far enough removed from an animal’s natural environment or behavioral repertoire, it may reflect neural or behavioral responses that are not used naturally (Krakauer et al., 2017).”

• cites Krakauer et al., 2017 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

Recent findings provide evidence of antisocial effects of OT.

“While seminal studies in male-male and male-female rodent pairs have contributed to the notion of OT as a facilitator of affiliative behaviors (Lukas et al., 2011, Williams et al., 1994, Witt et al., 1992), recent findings also provide evidence of antisocial effects of OT (van Anders et al., 2013, Bartz et al., 2011, Beery, 2015).”

• cites van Anders et al., 2013 sourceNot checked yet

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• cites Bartz et al., 2011Not checked yet

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• cites Beery, 2015 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

Seminal studies in male-male and male-female rodent pairs have contributed to the notion of OT as a facilitator of affiliative behaviors.

“While seminal studies in male-male and male-female rodent pairs have contributed to the notion of OT as a facilitator of affiliative behaviors (Lukas et al., 2011, Williams et al., 1994, Witt et al., 1992), recent findings also provide evidence of antisocial effects of OT (van Anders et al., 2013, Bartz et al., 2011, Beery, 2015).”

• cites Lukas et al., 2011 sourceNot checked yet

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• cites Williams et al., 1994 sourceNot checked yet

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• cites Witt et al., 1992 sourceNot checked yet

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Classified as probably not a substantial claim.

The social salience hypothesis explains contradictory effects of OT by suggesting that OT regulates the salience of social cues depending on contextual and individual variation.

“The social salience hypothesis explains these apparently contradictory effects by suggesting that OT acts to regulate the salience of social cues, depending on contextual and individual variation.”

• cites Shamay-Tsoory and Abu-Akel, 2016 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The receptor autoradiography procedure was performed according to Young et al., 1997.

“The receptor autoradiography procedure was performed according to Young et al., 1997 using a linear OT receptor antagonist [125I]-d(CH2)5[Tyr(Me)2-Tyr-Nh2]9-OVT (Perkin Elmer, USA) as a tracer.”

• cites Young et al., 1997 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

To exclude exhaustion of OT neurons or depletion of neuropeptide content, another set of operated mice was subjected to a regime of photoactivations similar to the main experiments.

“To exclude exhaustion of the OT neurons or depletion of neuropeptide content, another set of operated mice (Oxt::Cre+/− and Oxt::Cre−/−) were subjected to a regime of photoactivations similar to the main experiments (2 s long light pulses every 2 hr during the dark phase of the day, summing up to five activations per day).”

Probably not a substantial claim: may not pass S3 (load-bearing).

OT is being explored as a therapeutic agent in human disorders affecting social behavior and cognition, such as social anxiety and ASD.

“Since OT is being explored as a therapeutic agent in human disorders affecting social behavior and cognition, such as social anxiety and ASD (Alvares et al., 2017, Guastella et al., 2009, Ooi et al., 2017), these findings are crucial to consider.”

• cites Alvares et al., 2017 sourceNot checked yet

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• cites Guastella et al., 2009 sourceNot checked yet

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• cites Ooi et al., 2017Not checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The recombined regions of the BAC were sequence-verified.

“The recombined regions of the BAC were also sequence-verified.”

Probably not a substantial claim: may not pass S3 (load-bearing).

In rare cases where unusual tissue damage was observed in post-mortem IHC, mice were excluded from analysis.

“In rare cases, where unusual tissue damage was observed in post-mortem IHC, mice were excluded from analysis.”

Classified as probably not a substantial claim.

Plasma OT levels were measured using an extremely sensitive and specific RIA kit.

“Plasma was extracted and OT levels were measured using an extremely sensitive and specific RIA kit (RIAgnosis, Sinzing, Germany) (Jong et al., 2015, Kagerbauer et al., 2013).”

• cites Jong et al., 2015 sourceNot checked yet

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• cites Kagerbauer et al., 2013 sourceNot checked yet

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Probably not a substantial claim: may not pass S2 (checkable), S3 (load-bearing). May combine two claims.

This finding is in agreement with the social salience hypothesis and emphasizes the importance of observing behavioral changes over longer time periods and in a variety of contexts.

“This finding is in agreement with the social salience hypothesis and emphasizes the importance of observing behavioral changes over longer time periods, rather than only following acute manipulation, and in a variety of contexts.”

Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

Access resistance was continuously monitored and recordings were terminated if it changed by more than 15%.

“The access resistance (Ra) was continuously monitored and recordings were terminated if Ra changed > 15%.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Before each experiment, a 20 s pulse of 561 nm light was applied to convert all SSFO channels to the dark state and prevent run-down of photocurrents.

“Before each experiment, a 20 s pulse of 561 nm light was applied to convert all of the SSFO channels to the dark state and prevent run-down of the photocurrents.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Brain tissue damage was examined post-mortem and in most cases found to be as expected in cannulation experiments.

“Additionally, brain tissue damage was examined post-mortem and in most cases found to be as expected in cannulation experiments.”

Probably not a substantial claim: may not pass S3 (load-bearing).

In the rare occasion of inflammation, mice were excluded from further research.

“In the rare occasion of inflammation, mice were excluded from further research.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Early studies in the arenas showed that mice would spend on average 1-2 hr in the feeder zones when food was freely available.

“This time frame was chosen based on data from early studies in the arenas that showed mice would spend on average 1-2 hr in the feeder zones when food was freely available (Shemesh et al., 2013).”

• cites Shemesh et al., 2013 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing).

The alpha level of significance was adjusted using the Bonferroni method.

“Alpha level of significance was adjusted using the Bonferroni method.”

Probably not a substantial claim: may not pass S3 (load-bearing).

For electrophysiological data and grooming-related measurements in the group experiment, a one-tailed approach was employed.

“In electrophysiological data, as well as grooming-related measurements in the group experiment, a specific direction of the response was expected based on previous evidence, therefore a one-tailed approach was employed.”

• AssumptionRests on an assumption: A specific direction of the response was expected based on previous evidence for electrophysiological data and grooming-related measurements in the group experiment.

Probably not a substantial claim: may not pass S3 (load-bearing).

Frames selected for close social investigation were manually classified by human observers who were blind to the genotype of the group.

“These frames were then manually classified by human observers, who were blind to the genotype of the group, as either ‘face-to-face’ investigation, ‘face-to-back’ investigation, or ‘others’ (e.g., tail-to-tail contact).”

Probably not a substantial claim: may not pass S3 (load-bearing).

The previously-described automated tracking system is currently unable to automatically recognize specific body parts.

“Since our previously-described automated tracking system is currently unable to automatically recognize specific body parts, this system was only used to select from each video 150 frames in which exactly 2 mice were in close contact (< 3.5 cm between their body ends) in the open area.”

Probably not a substantial claim: may not pass S3 (load-bearing).

Introducing a female to the cage for 6 days and not changing the bedding improves the establishment of territory in male mice.

“To improve the establishment of territory, a female was introduced to the cages for 6 days and the bedding was not changed (Koolhaas et al., 2013).”

• cites Koolhaas et al., 2013 sourceNot checked yet

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Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

The data did not fulfil criteria for parametric testing (normality of residuals and equal variances between experimental groups), so non-parametric tests were employed.

“Non-parametric tests were employed, as data did not fulfil criteria for parametric testing (normality of residuals and equal variances between experimental groups).”

Probably not a substantial claim: may not pass S3 (load-bearing).

Trained observers who scored self-grooming were blind to the genotype of the mice.

“Trained observers, blind to the genotype of the mice, observed the recorded movies for a period of 5 min following each optogenetic activation.”

Classified as probably not a substantial claim.

Behavioral scoring in the resident-intruder test was performed by a trained observer blind to the treatment.

“Behavioral scoring was performed by a trained observer blind to the treatment using the ‘Solomon Coder’ freeware (https://solomon.andraspeter.com/).”