A novel naïve Bayes approach to identifying grooming behaviors in the force-plate actometric platform.

BACKGROUND: Self-grooming behavior in rodents serves as a valuable behavioral index for investigating stereotyped and perseverative responses. Most current grooming analyses rely on video observation, which lacks standardization, efficiency, and quantitative information about force. To address these limitations, we developed an automated paradigm to analyze grooming using a force-plate actometer. NEW METHOD: Grooming behavior is quantified by calculating ratios of relevant movement power spectral bands. These ratios are input into a naïve Bayes classifier, trained with manual video observations. The effectiveness of this method was tested using CIN-d mice, an animal model developed through early-life depletion of striatal cholinergic interneurons (CIN-d) and featuring prolonged grooming responses to acute stressors. Behavioral monitoring was simultaneously conducted on the force-place actometer and by video recording. RESULTS: The naïve Bayes approach achieved 93.7% accurate classification and an area under the receiver operating characteristic curve of 0.894. We confirmed that male CIN-d mice displayed significantly longer grooming durations than controls. However, this elevation was not correlated with increases in grooming force. Notably, the dopaminergic antagonist haloperidol reduced grooming force and duration. COMPARISON WITH EXISTING METHODS: In contrast to observation-based approaches, our method affords rapid, unbiased, and automated assessment of grooming duration, frequency, and force. CONCLUSIONS: Our novel approach enables fast and accurate automated detection of grooming behaviors. This method holds promise for high-throughput assessments of grooming stereotypies in animal models of neuropsychiatric disorders.

A novel naïve Bayes approach to identifying grooming behaviors in the force-plate actometric platform.

Background: Self-grooming behavior in rodents serves as a valuable model for investigating stereotyped and perseverative responses. Most current grooming analyses primarily rely on video observation, which lacks standardization, efficiency, and quantitative information about force. To address these limitations, we developed an automated paradigm to analyze grooming using a force-plate actometer. New Method: Grooming behavior is quantified by calculating ratios of relevant movement power spectral bands. These ratios are then input into a naïve Bayes classifier, trained with manual video observations. To validate the effectiveness of this method, we applied it to the behavioral analysis of the early-life striatal cholinergic interneuron depletion (CIN-d) mouse, a model of tic pathophysiology recently developed in our laboratory, which exhibits prolonged grooming responses to acute stressors. Behavioral monitoring was simultaneously conducted on the force-place actometer and by video recording. Results: The naïve Bayes approach achieved 93.7% accurate classification and an area under the receiver operating characteristic curve of 0.894. We confirmed that male CIN-d mice displayed significantly longer grooming durations compared to controls. However, this elevation was not correlated with increases in grooming force. Notably, haloperidol, a benchmark therapy for tic disorders, reduced both grooming force and duration. Comparison with Existing Methods: In contrast to observation-based approaches, our method affords rapid, unbiased, and automated assessment of grooming duration, frequency, and force. Conclusions: Our novel approach enables fast and accurate automated detection of grooming behaviors. This method holds promise for high-throughput assessments of grooming stereotypies in animal models of tic disorders and other psychiatric conditions.

Prefrontal allopregnanolone mediates the adverse effects of acute stress in a mouse model of tic pathophysiology.

Ample evidence suggests that acute stress can worsen symptom severity in Tourette syndrome (TS); however, the neurobiological underpinnings of this phenomenon remain poorly understood. We previously showed that acute stress exacerbates tic-like and other TS-associated responses via the neurosteroid allopregnanolone (AP) in an animal model of repetitive behavioral pathology. To verify the relevance of this mechanism to tic pathophysiology, here we tested the effects of AP in a mouse model recapitulating the partial depletion of dorsolateral cholinergic interneurons (CINs) seen in post-mortem studies of TS. Mice underwent targeted depletion of striatal CINs during adolescence and were tested in young adulthood. Compared with controls, partially CIN-depleted male mice exhibited several TS-relevant abnormalities, including deficient prepulse inhibition (PPI) and increased grooming stereotypies after a 30-min session of spatial confinement - a mild acute stressor that increases AP levels in the prefrontal cortex (PFC). These effects were not seen in females. Systemic and intra-PFC AP administration dose-dependently worsened grooming stereotypies and PPI deficits in partially CIN-depleted males. Conversely, both AP synthesis inhibition and pharmacological antagonism reduced the effects of stress. These results further suggest that AP in the PFC mediates the adverse effects of stress on the severity of tics and other TS-related manifestations. Future studies will be necessary to confirm these mechanisms in patients and define the circuitry responsible for the effects of AP on tics.

Slc9a6 mutation causes Purkinje cell loss and ataxia in the shaker rat.

The shaker rat carries a naturally occurring mutation leading to progressive ataxia characterized by Purkinje cell (PC) loss. We previously reported on fine-mapping the shaker locus to the long arm of the rat X chromosome. In this work, we sought to identify the mutated gene underlying the shaker phenotype and confirm its identity by functional complementation. We fine-mapped the candidate region and analyzed cerebellar transcriptomes, identifying a XM_217630.9 (Slc9a6):c.[191_195delinsA] variant in the Slc9a6 gene that segregated with disease. We generated an adeno-associated virus (AAV) targeting Slc9a6 expression to PCs using the mouse L7-6 (L7) promoter. We administered the AAV prior to the onset of PC degeneration through intracerebroventricular injection and found that it reduced the shaker motor, molecular and cellular phenotypes. Therefore, Slc9a6 is mutated in shaker and AAV-based gene therapy may be a viable therapeutic strategy for Christianson syndrome, also caused by Slc9a6 mutation.

Computational investigation of the impact of deep brain stimulation contact size and shape on neural selectivity.

Background.Understanding neural selectivity is essential for optimizing medical applications of deep brain stimulation (DBS). We previously showed that modulation of the DBS waveform can induce changes in orientation-based selectivity, and that lengthening of DBS pulses or directional segmentation can reduce preferential selectivity for large axons. In this work, we sought to investigate a simple, but important question from a generalized perspective: how do the size and shape of the contact influence neural selectivity?Methods.We created multicompartment neuron models for several axon diameters and used finite element modeling with standard-sized cylindrical leads to determine the effects on changing contact size and shape on axon activation profiles and volumes of tissue activated. Contacts ranged in size from 0.04 to 16 mm2, compared with a standard size of 6 mm2.Results.We found that changes in contact size are predicted to induce substantial changes in orientation-based selectivity in the context of a cylindrical lead, and changes in contact width or height can alter this selectivity. Smaller contact sizes were more effective in constraining neural activation to small, nearby axons. However, micro-scale contacts enable only limited spread of neural activation before exceeding standard charge density limitations; further, energetic efficiency is optimized by somewhat larger contacts.Interpretations.Small-scale contacts may be optimal for constraining stimulation in nearby grey matter and avoiding orientation-selective activation. However, given charge density limitations and energy inefficiency of micro-scale contacts, we predict that contacts sized similarly to or slightly smaller than segmented clinical leads may optimize energy efficiency while avoiding charge density limitations.

Detecting and Quantifying Ataxia-Related Motor Impairments in Rodents Using Markerless Motion Tracking With Deep Neural Networks.

In this study we evaluate the application of video-based markerless motion tracking based on deep neural networks for the analysis of ataxia-specific movement abnormalities in rodent models of cerebellar ataxia. Based on a small amount (<100) of manually labeled video frames, markerless motion tracking enabled the extraction of movement trajectories and parameters characterizing ataxia-specific movement abnormalities. In the first experiment, we analyzed videos of 6 shaker and 4 wildtype rats and were able to reproduce thê5 Hz tremor frequency in the shaker rat without the usage of a force plate. In the second experiment, we investigated a spinocerebellar ataxia type 3 (SCA3) mouse model (6 mice aged 3 months and 3 mice aged 9 months) in a beam-balancing task. By establishing a parameter for the assessment of rhythmicity of gait (RoG), we not only found a significantly higher RoG in wildtype mice compared to affected SCA3 mice aged 9 months, but were also able to reveal a significantly lower than typical RoG in SCA3 mice aged 3 months which exhibit no abnormalities in visual inspection. These prototypical results suggest the capability of the presented methods for the application in upcoming therapeutic intervention trials to identify subtle changes in movement behavior.

Neural selectivity, efficiency, and dose equivalence in deep brain stimulation through pulse width tuning and segmented electrodes.

BACKGROUND: Achieving deep brain stimulation (DBS) dose equivalence is challenging, especially with pulse width tuning and directional contacts. Further, the precise effects of pulse width tuning are unknown, and recent reports of the effects of pulse width tuning on neural selectivity are at odds with classic biophysical studies. METHODS: We created multicompartment neuron models for two axon diameters and used finite element modeling to determine extracellular influence from standard and segmented electrodes. We analyzed axon activation profiles and calculated volumes of tissue activated. RESULTS: We find that long pulse widths focus the stimulation effect on small, nearby fibers, suppressing distant white matter tract activation (responsible for some DBS side effects) and improving battery utilization when equivalent activation is maintained for small axons. Directional leads enable similar benefits to a greater degree. Reexamining previous reports of short pulse stimulation reducing side effects, we explore a possible alternate explanation: non-dose equivalent stimulation may have resulted in reduced spread of neural activation. Finally, using internal capsule avoidance as an example in the context of subthalamic stimulation, we present a patient-specific model to show how long pulse widths could help increase the biophysical therapeutic window. DISCUSSION: We find agreement with classic studies and predict that long pulse widths may focus the stimulation effect on small, nearby fibers and improve power consumption. While future pre-clinical and clinical work is necessary regarding pulse width tuning, it is clear that future studies must ensure dose equivalence, noting that energy- and charge-equivalent amplitudes do not result in equivalent spread of neural activation when changing pulse width.

Current Opinions and Consensus for Studying Tremor in Animal Models.

Tremor is the most common movement disorder; however, we are just beginning to understand the brain circuitry that generates tremor. Various neuroimaging, neuropathological, and physiological studies in human tremor disorders have been performed to further our knowledge of tremor. But, the causal relationship between these observations and tremor is usually difficult to establish and detailed mechanisms are not sufficiently studied. To overcome these obstacles, animal models can provide an important means to look into human tremor disorders. In this manuscript, we will discuss the use of different species of animals (mice, rats, fruit flies, pigs, and monkeys) to model human tremor disorders. Several ways to manipulate the brain circuitry and physiology in these animal models (pharmacology, genetics, and lesioning) will also be discussed. Finally, we will discuss how these animal models can help us to gain knowledge of the pathophysiology of human tremor disorders, which could serve as a platform towards developing novel therapies for tremor.

Deep cerebellar stimulation reduces ataxic motor symptoms in the shaker rat.

OBJECTIVE: Degenerative cerebellar ataxias (DCAs) affect up to 1 in 5,000 people worldwide, leading to incoordination, tremor, and falls. Loss of Purkinje cells, nearly universal across DCAs, dysregulates the dentatothalamocortical network. To address the paucity of treatment strategies, we developed an electrical stimulation-based therapy for DCAs targeting the dorsal dentate nucleus. METHODS: We tested this therapeutic strategy in the Wistar Furth shaker rat model of Purkinje cell loss resulting in tremor and ataxia. We implanted shaker rats with stimulating electrodes targeted to the dorsal dentate nucleus and tested a spectrum of frequencies ranging from 4 to 180 Hz. RESULTS: Stimulation at 30 Hz most effectively reduced motor symptoms. Stimulation frequencies >100 Hz, commonly used for parkinsonism and essential tremor, worsened incoordination, and frequencies within the tremor physiologic range may worsen tremor. INTERPRETATION: Low-frequency deep cerebellar stimulation may provide a novel strategy for treating motor symptoms of degenerative cerebellar ataxias. Ann Neurol 2019;85:681-690.

Deep brain stimulation exacerbates hypokinetic dysarthria in a rat model of Parkinson's disease.

Motor symptoms of Parkinson's disease (PD) follow the degeneration of dopaminergic neurons in the substantia nigra pars compacta. Deep brain stimulation (DBS) treats some parkinsonian symptoms, such as tremor, rigidity, and bradykinesia, but may worsen certain medial motor symptoms, including hypokinetic dysarthria. The mechanisms by which DBS exacerbates dysarthria while improving other symptoms are unclear and difficult to study in human patients. This study proposes an animal model of DBS-exacerbated dysarthria. We use the unilateral, 6-hydroxydopamine (6-OHDA) rat model of PD to test the hypothesis that DBS exacerbates quantifiable aspects of vocalization. Mating calls were recorded from sexually experienced male rats under healthy and parkinsonian conditions and during DBS of the subthalamic nucleus. Relative to healthy rats, parkinsonian animals made fewer calls with shorter and less complex vocalizations. In the parkinsonian rats, putatively therapeutic DBS further reduced call frequency, duration, and complexity. The individual utterances of parkinsonian rats spanned a greater bandwidth than those of healthy rats, potentially reducing the effectiveness of the vocal signal. This utterance bandwidth was further increased by DBS. We propose that the parkinsonism-associated changes in call frequency, duration, complexity, and dynamic range combine to constitute a rat analog of parkinsonian dysarthria. Because DBS exacerbates the parkinsonism-associated changes in each of these metrics, the subthalamic stimulated 6-OHDA rat is a good model of DBS-induced hypokinetic dysarthria in PD. This model will help researchers examine how DBS alleviates many motor symptoms of PD while exacerbating parkinsonian speech deficits that can greatly diminish patient quality of life.

Subthalamic deep brain stimulation reduces pathological information transmission to the thalamus in a rat model of parkinsonism.

The degeneration of dopaminergic neurons in the substantia nigra pars compacta leads to parkinsonian motor symptoms via changes in electrophysiological activity throughout the basal ganglia. High-frequency deep brain stimulation (DBS) partially treats these symptoms, but the mechanisms are unclear. We hypothesize that motor symptoms of Parkinson's disease (PD) are associated with increased information transmission from basal ganglia output neurons to motor thalamus input neurons and that therapeutic DBS of the subthalamic nucleus (STN) treats these symptoms by reducing this extraneous information transmission. We tested these hypotheses in a unilateral, 6-hydroxydopamine-lesioned rodent model of hemiparkinsonism. Information transfer between basal ganglia output neurons and motor thalamus input neurons increased in both the orthodromic and antidromic directions with hemiparkinsonian (hPD) onset, and these changes were reversed by behaviorally therapeutic STN-DBS. Omnidirectional information increases in the parkinsonian state underscore the detrimental nature of that pathological information and suggest a loss of information channel independence. Therapeutic STN-DBS reduced that pathological information, suggesting an effective increase in the number of independent information channels. We interpret these data with a model in which pathological information and fewer information channels diminishes the scope of possible motor activities, driving parkinsonian symptoms. In this model, STN-DBS restores information-channel independence by eliminating or masking the parkinsonism-associated information, and thus enlarges the scope of possible motor activities, alleviating parkinsonian symptoms.

Telomere length is a determinant of emphysema susceptibility.

RATIONALE: Germline mutations in the enzyme telomerase cause telomere shortening, and have their most common clinical manifestation in age-related lung disease that manifests as idiopathic pulmonary fibrosis. Short telomeres are also a unique heritable trait that is acquired with age. OBJECTIVES: We sought to understand the mechanisms by which telomerase deficiency contributes to lung disease. METHODS: We studied telomerase null mice with short telomeres. MEASUREMENTS AND MAIN RESULTS: Although they have no baseline histologic defects, when mice with short telomeres are exposed to chronic cigarette smoke, in contrast with controls, they develop emphysematous air space enlargement. The emphysema susceptibility did not depend on circulating cell genotype, because mice with short telomeres developed emphysema even when transplanted with wild-type bone marrow. In lung epithelium, cigarette smoke exposure caused additive DNA damage to telomere dysfunction, which limited their proliferative recovery, and coincided with a failure to down-regulate p21, a mediator of cellular senescence, and we show here, a determinant of alveolar epithelial cell cycle progression. We also report early onset of emphysema, in addition to pulmonary fibrosis, in a family with a germline deletion in the Box H domain of the RNA component of telomerase. CONCLUSIONS: Our data indicate that short telomeres lower the threshold of cigarette smoke-induced damage, and implicate telomere length as a genetic susceptibility factor in emphysema, potentially contributing to its age-related onset in humans.

Ancestral mutation in telomerase causes defects in repeat addition processivity and manifests as familial pulmonary fibrosis.

The telomerase reverse transcriptase synthesizes new telomeres onto chromosome ends by copying from a short template within its integral RNA component. During telomere synthesis, telomerase adds multiple short DNA repeats successively, a property known as repeat addition processivity. However, the consequences of defects in processivity on telomere length maintenance are not fully known. Germline mutations in telomerase cause haploinsufficiency in syndromes of telomere shortening, which most commonly manifest in the age-related disease idiopathic pulmonary fibrosis. We identified two pulmonary fibrosis families that share two non-synonymous substitutions in the catalytic domain of the telomerase reverse transcriptase gene hTERT: V791I and V867M. The two variants fell on the same hTERT allele and were associated with telomere shortening. Genealogy suggested that the pedigrees shared a single ancestor from the nineteenth century, and genetic studies confirmed the two families had a common founder. Functional studies indicated that, although the double mutant did not dramatically affect first repeat addition, hTERT V791I-V867M showed severe defects in telomere repeat addition processivity in vitro. Our data identify an ancestral mutation in telomerase with a novel loss-of-function mechanism. They indicate that telomere repeat addition processivity is a critical determinant of telomere length and telomere-mediated disease.