The neurophysiological potential:Translatability and reproducibility of preclinical outcome measures in psychiatric research

Animal models are an important source of information for studying the biological mechanisms and complexity of mental disorders. In this thesis we studied which characteristics are most suitable to study psychiatric disorders in animal models. We sought for characteristics of which the findings in animals apply to patients and which are robust enough to give the same results when tested in different laboratories. Behavioural changes are important in psychiatric disorders, but turn out to be difficult to study in animal models. In a study where we combined all available data on the behaviour of an autism mouse mode, we found that the results were highly inconsistent, although mostly representing the symptoms of patients well. Physiological characteristics have the potential to generate more valuable results than behaviour, amongst other things because they can be measured in patients and animals in more similar ways. In the same mouse model, we studied brain activity in rest and in response to sensory stimulation. The responses to sensory stimuli also showed inconsistent results, however the characteristics of the brain activity in rest were consistent and in line with earlier findings. The brain activity of the mice was dominated by inhibitory activity. This was in contrast with the hypothesis of a hyperactive brain in autism, but in line with studies assessing the brain activity of autistic children using the same method. Our results suggest that studying physiological characteristics in psychiatric disorders can lead to more robust findings which translate better between the animal models and patients

Largely unaffected auditory and visual sensory processing phenotypes in the evoked potentials of Fmr1 KO2 mice

Sensory sensitivity symptoms are common in autism spectrum disorders and fragile X syndrome. Mainly in the auditory modality, disturbed processing has been found in both fragile X patients and the corresponding genetic mouse model, the Fmr1 knockout mouse. Here, we tried to replicate the auditory deficits and assess whether also visual processing is affected, using electroencephalography readouts under freely behaving conditions in the second-generation Fmr1 knockout mice. No differences between wild-type and knockout animals were found in single auditory and visual evoked potentials in response to pure sine tones and full-field light flashes. Visual sensory gating was enhanced in the early but not the late components of the evoked potentials, but no changes were found in auditory sensory gating. The higher harmonics of the synchronisation response to flickering visual stimuli seemed to be reduced with 10, but not 20 or 40 Hz, stimulation. However, this effect was not reproduced in an independent second cohort of animals. No synchronisation differences were found in response to a chirp stimulus, of which the frequency steadily increased. Taken together, this study could not reproduce earlier reported increased amplitudes in auditory responses, nor could it convincingly show that synchronisation deficits found to be present in the auditory modality also existed in the visual modality. The discrepancies within this study as well as between various studies assessing sensory processing in the Fmr1 KO raise questions about the external validity of these phenotypes and warrant careful interpretation of these phenotypes

Translational validity and methodological underreporting in animal research:A systematic review and meta-analysis of the Fragile X syndrome (Fmr1 KO) rodent model

Predictive models are essential for advancing knowledge of brain disorders. High variation in study outcomes hampers progress. To address the validity of predictive models, we performed a systematic review and meta-analysis on behavioural phenotypes of the knock-out rodent model for Fragile X syndrome according to the PRISMA reporting guidelines. In addition, factors accountable for the heterogeneity between findings were analyzed. The knock-out model showed good translational validity and replicability for hyperactivity, cognitive and seizure phenotypes. Despite low replicability, translational validity was also found for social behaviour and sensory sensitivity, but not for attention, aggression and cognitive flexibility. Anxiety, acoustic startle and prepulse inhibition phenotypes, despite low replicability, were opposite to patient symptomatology. Subgroup analyses for experimental factors moderately explain the low replicability, these analyses were hindered by under-reporting of methodologies and environmental conditions. Together, the model has translational validity for most clinical phenotypes, but caution must be taken due to low effect sizes and high inter-study variability. These findings should be considered in view of other rodent models in preclinical research

Unaffected sensory processing in FMR1 KO mice

Sensory sensitivity problems are a common symptom in autism spectrum disorders and fragile X syndrome. Mainly in the auditory modality, disturbed sensory processing has been found in both fragile-X patients as well as the knockout animals of the corresponding genetic model, the Fmr1 knockout mouse. Here we assessed not only auditory but also visual sensory processing in the second generation Fmr1 KO mice (Mientjes et al., 2006) under freely behaving conditions using electroencephalography readouts. No differences between wildtype and knockout animals were found in single auditory and visual evoked potentials in response to pure sine tones and full-field white light flashes. Visual sensory gating was enhanced in the early but not the late components of the evoked potentials, no changes were found in auditory gating. Synchronization to a steady state flickering visual stimulus seemed to be reduced in the higher harmonic responses of 10Hz, but not 20 or 40Hz, stimulation. However, this effect could not be reproduced in an independent cohort of animals. No differences were found in the intertrial phase consistency, nor in power in response to a chirp stimulus in which the frequency steadily increased. This study could not reproduce earlier found increased amplitudes in auditory stimuli, nor could it convincingly show that synchronization deficits found to be present in response to auditory stimuli also existed in the visual modality. Taken together, no robust auditory and visual processing deficits could be found in the Fmr1 knockout mice. When aiming to study sensory processing in the Fmr1 knockout mice, the appropriate paradigms should be carefully selected to improve translatability and predictive validity

Neuronal Oscillation Dynamics Fmr1 KO2 mice

In vitro and ex vivo studies have shown consistent indications of hyperexcitability in the Fmr1 KO mouse model of autism spectrum disorder. We recently introduced a method to quantify network-level functional E/I ratio (fEI) from the neuronal oscillations. Here, we used this measure to study whether the implicated synaptic E/I disturbances translate to disturbances in network physiology in the Fmr1 KO model. We performed wireless skull-EEG recordings in wildtype (WT) and knockout (KO) male mice of the Fmr1 KO2 line in two separate experiments as an internal replication. Vigilance-state scoring was used to extract segments of inactive wakefulness as an equivalent behavioural condition to the human resting-state and, subsequently, we performed high-frequency resolution analysis of the fE/I biomarker, long-range temporal correlations (LRTC), and spectral power. We corroborated earlier studies showing increased high-frequency power in Fmr1 KO mice. LRTCs were higher in the gamma frequency ranges. Contrary to expectations, fEI was lower in the KO mice in high frequency ranges, suggesting more inhibition-dominated networks. Exposure to the GABA-agonist clonazepam decreased the fEI in both genotypes, confirming that increasing inhibitory tone results in a reduction of fEI. In addition, clonazepam decreased EEG power and increased LRTCs. These findings show applicability of these new resting-state EEG biomarkers to animal models for translational studies. They allow investigation of the effects of lower-level disturbances in E/I balance in relation to the organization of mass brain activity and how these may result in counterintuitive dynamics

Neuronal Oscillation Dynamics Fmr1 KO2 mice

In vitro and ex vivo studies have shown consistent indications of hyperexcitability in the Fmr1 KO mouse model of autism spectrum disorder. We recently introduced a method to quantify network-level functional E/I ratio (fEI) from the neuronal oscillations. Here, we used this measure to study whether the implicated synaptic E/I disturbances translate to disturbances in network physiology in the Fmr1 KO model. We performed wireless skull-EEG recordings in wildtype (WT) and knockout (KO) male mice of the Fmr1 KO2 line in two separate experiments as an internal replication. Vigilance-state scoring was used to extract segments of inactive wakefulness as an equivalent behavioural condition to the human resting-state and, subsequently, we performed high-frequency resolution analysis of the fE/I biomarker, long-range temporal correlations (LRTC), and spectral power. We corroborated earlier studies showing increased high-frequency power in Fmr1 KO mice. LRTCs were higher in the gamma frequency ranges. Contrary to expectations, fEI was lower in the KO mice in high frequency ranges, suggesting more inhibition-dominated networks. Exposure to the GABA-agonist clonazepam decreased the fEI in both genotypes, confirming that increasing inhibitory tone results in a reduction of fEI. In addition, clonazepam decreased EEG power and increased LRTCs. These findings show applicability of these new resting-state EEG biomarkers to animal models for translational studies. They allow investigation of the effects of lower-level disturbances in E/I balance in relation to the organization of mass brain activity and how these may result in counterintuitive dynamics

Neuronal Oscillation Dynamics Fmr1 KO2 mice

In vitro and ex vivo studies have shown consistent indications of hyperexcitability in the Fmr1 KO mouse model of autism spectrum disorder. We recently introduced a method to quantify network-level functional E/I ratio (fEI) from the neuronal oscillations. Here, we used this measure to study whether the implicated synaptic E/I disturbances translate to disturbances in network physiology in the Fmr1 KO model. We performed wireless skull-EEG recordings in wildtype (WT) and knockout (KO) male mice of the Fmr1 KO2 line in two separate experiments as an internal replication. Vigilance-state scoring was used to extract segments of inactive wakefulness as an equivalent behavioural condition to the human resting-state and, subsequently, we performed high-frequency resolution analysis of the fE/I biomarker, long-range temporal correlations (LRTC), and spectral power. We corroborated earlier studies showing increased high-frequency power in Fmr1 KO mice. LRTCs were higher in the gamma frequency ranges. Contrary to expectations, fEI was lower in the KO mice in high frequency ranges, suggesting more inhibition-dominated networks. Exposure to the GABA-agonist clonazepam decreased the fEI in both genotypes, confirming that increasing inhibitory tone results in a reduction of fEI. In addition, clonazepam decreased EEG power and increased LRTCs. These findings show applicability of these new resting-state EEG biomarkers to animal models for translational studies. They allow investigation of the effects of lower-level disturbances in E/I balance in relation to the organization of mass brain activity and how these may result in counterintuitive dynamics

Neuronal Oscillation Dynamics Fmr1 KO2 mice

In vitro and ex vivo studies have shown consistent indications of hyperexcitability in the Fmr1 KO mouse model of autism spectrum disorder. We recently introduced a method to quantify network-level functional E/I ratio (fEI) from the neuronal oscillations. Here, we used this measure to study whether the implicated synaptic E/I disturbances translate to disturbances in network physiology in the Fmr1 KO model. We performed wireless skull-EEG recordings in wildtype (WT) and knockout (KO) male mice of the Fmr1 KO2 line in two separate experiments as an internal replication. Vigilance-state scoring was used to extract segments of inactive wakefulness as an equivalent behavioural condition to the human resting-state and, subsequently, we performed high-frequency resolution analysis of the fE/I biomarker, long-range temporal correlations (LRTC), and spectral power. We corroborated earlier studies showing increased high-frequency power in Fmr1 KO mice. LRTCs were higher in the gamma frequency ranges. Contrary to expectations, fEI was lower in the KO mice in high frequency ranges, suggesting more inhibition-dominated networks. Exposure to the GABA-agonist clonazepam decreased the fEI in both genotypes, confirming that increasing inhibitory tone results in a reduction of fEI. In addition, clonazepam decreased EEG power and increased LRTCs. These findings show applicability of these new resting-state EEG biomarkers to animal models for translational studies. They allow investigation of the effects of lower-level disturbances in E/I balance in relation to the organization of mass brain activity and how these may result in counterintuitive dynamics