Cynthia A. Kelm-Nelson

Parkinson's disease is a progressive neurodegenerative disorder with an unclear etiology and a long preclinical phase. The multitude of challenges of studying this in humans highlight the need for robust preclinical animal models that accurately reflect the disease's preclinical progression. While traditional neurotoxin models fail to capture the slow, genetic nature of PD, genetic models, such as the phosphatase and tensin homolog-induced putative kinase 1 (Pink1-/-) knockout rat, offer a preclinical, translational approach. This review synthesizes the current understanding of the Pink1-/- rat model, focusing on two key aspects: neuroinflammation and vocalization deficits. The Pink1-/- rat presents early non-motor symptoms, including vocalization deficits, which can serve as a non-invasive, objective, and longitudinal biomarker of disease progression. This review further proposes a direct link between the model's neuroinflammatory profile, characterized by chronic glial activation and the upregulation of inflammatory pathways, and the observed vocal dysfunctions. By bridging these two critical areas, neuroinflammation as a pathological mechanism and vocalizations as a functional readout, this review provides a framework for future investigations into the diagnosis and treatment of Parkinson’s disease and suggests that targeting specific inflammatory pathways could be a viable therapeutic strategy.

Introduction: Ultrasonic vocalization (USV) can indicate affective states—including psychosocial stress—in mice and rats. However, stress-induced USV changes could be confounded by laboratory experimental variables such as the type of behavioral stress paradigm, the elicitation method, rodent strain, etc. We sought to provide a review of the current literature to delineate how psychosocial stress-altered rodent USVs may be affected by factors of age, sex, strain, species, elicitation paradigm, and stressor. Methods: We used PubMed, Scopus (Elsevier), PsycINFO (EBSCO), and the following Web of Science (Clarivate) databases: Biological Abstracts, CAB Abstracts, Science Citation Index-Expanded, and Emerging Sources Citation Index. The studies identified by our search strategy were independently screened by two authors with the following inclusion criteria: peer-reviewed, in English, reported original data, and described USV in response to stress in rats or mice. The data extracted included USV acoustic parameters (mean peak frequency and mean amplitude (loudness)), details of the stress and USV elicitation paradigms, rodent species, age, and sex variables. Results: The following screening of 5309 titles/abstracts and 687 full-text articles revealed 148 articles. Footshock (20%), cold exposure (14%), and maternal separation (23.5%) were the most commonly used stress paradigms (duration and type of stressor varied across studies), with the total number of USV calls being the most commonly reported acoustic outcome. In rats, 121 articles described stress-altered USVs, while 25 studies reported the same in mice, and two reported multiple rodent species (rats and mice, alongside other rodent species such as gerbils). With respect to stress-altered USV changes with age, mice and rats increase USV rates after birth, with a peak around 6 to 10 days, and decrease USVs until weanling age. Of the five studies that reported sex-related differences in stress-induced USVs, females had an increased number of calls and lower average peak frequency in response to stress when compared to males. Only two to four studies reported strain-related differences in stress-induced vocalizations in rats and mice, respectively. Conclusions: The data from this review lay the groundwork for better understanding rodent USVs in response to psychosocial stress with effects of elicitation paradigm, stressor, age, and sex.

Parkinson disease (PD) is the fastest growing neurodegenerative disease. The molecular pathology of PD in the prodromal phase is poorly understood; as such, there are no specific prognostic or diagnostic tests. A validated Pink1 genetic knockout rat was used to model early-onset and progressive PD. Male Pink1−/− rats exhibit progressive declines in ultrasonic vocalizations as well as hindlimb and forelimb motor deficits by mid-to-late adulthood. Previous RNA-sequencing work identified upregulation of genes involved in disease pathways and inflammation within the brainstem and vocal fold muscle. The purpose of this study was to identify gene pathways within the whole blood of young Pink1−/− rats (3 months of age) and to link gene expression to early acoustical changes. To accomplish this, limb motor testing (open field and cylinder tests) and ultrasonic vocalization data were collected, immediately followed by the collection of whole blood and RNA extraction. Illumina® Total RNA-Seq TruSeq platform was used to profile differential expression of genes. Statistically significant genes were identified and Weighted Gene Co-expression Network Analysis was used to construct co-expression networks and modules from the whole blood gene expression dataset as well as the open field, cylinder, and USV acoustical dataset. ENRICHR was used to identify the top up-regulated biological pathways. The data suggest that inflammation and interferon signaling upregulation in the whole blood is present during early PD. We also identified genes involved in the dysregulation of ribosomal protein and RNA processing gene expression as well as prion protein gene expression. These data identified several potential blood biomarkers and pathways that may be linked to anxiety and vocalization acoustic parameters and are key candidates for future drug-repurposing work and comparison to human datasets.

Alzheimer’s disease (AD) is a complex neurodegenerative disorder with both genetic and non-genetic causes. Animal research models are available for a multitude of diseases and conditions affecting the central nervous system (CNS), and large-scale CNS gene expression data exist for many of these. Although there are several models specifically for AD, each recapitulates different aspects of the human disease. In this study we evaluate over 500 animal models to identify those with CNS gene expression patterns matching human AD datasets. Approaches included a hypergeometric based scoring system that rewards congruent gene expression patterns but penalizes discordant gene expression patterns. The top two models identified were APP/PS1 transgenic mice expressing mutant APP and PSEN1, and mice carrying a GFAP mutation that is causative of Alexander disease, a primary disorder of astrocytes in the CNS. The APP/PS1 and GFAP models both matched over 500 genes moving in the same direction as in human AD, and both had elevated GFAP expression and were highly congruent with one another. Also scoring highly were the 5XFAD model (with five mutations in APP and PSEN1) and mice carrying CK-p25, APP, and MAPT mutations. Animals with the APOE3 and 4 mutations combined with traumatic brain injury ranked highly. Bulbectomized rats scored high, suggesting anosmia could be causative of AD-like gene expression. Other matching models included the SOD1G93A strain and knockouts for SNORD116 (Prader-Willi mutation), GRID2, INSM1, XBP1, and CSTB. Many top models demonstrated increased expression of GFAP, and results were similar across multiple human AD datasets. Heatmap and Uniform Manifold Approximation Plot results were consistent with hypergeometric ranking. Finally, some gene manipulation models, including for TYROBP and ATG7, were identified with reversed AD patterns, suggesting possible neuroprotective effects. This study provides insight for the pathobiology of AD and the potential utility of available animal models.

Background Parkinson disease (PD) is the fastest growing neurodegenerative disease. The molecular pathology of PD in the prodromal phase is poorly understood; as such, there are no specific prognostic or diagnostic tests. A validated Pink1 genetic knockout rat was used to model early-onset and progressive PD. Male Pink1-/- rats exhibit progressive declines in ultrasonic vocalizations as well as hindlimb and forelimb motor deficits by mid-to-late adulthood. Previous RNA-sequencing work identified upregulation of genes involved in disease pathways and inflammation within the brainstem and vocal fold muscle. The purpose of this study was to identify gene pathways within the whole blood of young Pink1-/- rats (3 months of age) and to link gene expression to early acoustical changes. To accomplish this, limb motor testing (open field and cylinder tests) and ultrasonic vocalization data were collected, immediately followed by the collection of whole blood and RNA extraction. Illumina® Total RNA-Seq TruSeq platform was used to profile differential expression of genes. Statistically significant genes were identified and Weighted Gene Co-expression Network Analysis was used to construct co-expression networks and modules from the whole blood gene expression dataset as well as the open field, cylinder, and USV acoustical dataset. ENRICHR was used to identify the top up-regulated biological pathways. Results The data suggest that inflammation and interferon signaling upregulation in the whole blood is present during early PD. We also identified genes involved in the dysregulation of ribosomal protein and RNA processing gene expression as well as prion protein gene expression. Conclusions These data identified several potential blood biomarkers and pathways that may be linked to anxiety and vocalization acoustic parameters and are key candidates for future drug-repurposing work and comparison to human datasets.

Objectives and hypothesis: Vocal dysfunction, including hypophonia, in Parkinson disease (PD) manifests in the prodromal period and significantly impacts an individual's quality of life. Data from human studies suggest that pathology leading to vocal deficits may be structurally related to the larynx and its function. The Pink1-/- rat is a translational model used to study pathogenesis in the context of early-stage mitochondrial dysfunction. The primary objective of this work was to identify differentially expressed genes in the thyroarytenoid muscle and examine the dysregulated biological pathways in the female rat. Methods: RNA sequencing was used to determine thyroarytenoid (TA) muscle gene expression in adult female Pink1-/- rats compared with controls. A bioinformatic approach and the ENRICHR gene analysis tool were used to compare the sequencing dataset with biological pathways and processes, disease relationships, and drug-repurposing compounds. Weighted Gene Co-expression Network Analysis was used to construct biological network modules. The data were compared with a previously published dataset in male rats. Results: Significant upregulated pathways in female Pink1-/- rats included fatty acid oxidation and muscle contraction, synaptic transmission, and neuromuscular processes. Downregulated pathways included anterograde transsynaptic signaling, chemical synaptic transmission, and ion release. Several drug treatment options including cetuximab, fluoxetine, and resveratrol are hypothesized to reverse observed genetic dysregulation. Conclusions: Data presented here are useful for identifying biological pathways that may underlie the mechanisms of peripheral dysfunction including neuromuscular synaptic transmission to the TA muscle. These experimental biomarkers have the potential to be targeted as sites for improving the treatment for hypophonia in early-stage PD.

Parkinson’s disease (PD) is a progressive, degenerative disease that affects nearly 10 million people worldwide. Hallmark limb motor signs and dopamine depletion have been well studied; however, few studies evaluating early stage, prodromal biology exist. Pink1−/− rats, a rodent model of PD mitochondrial dysfunction, exhibit early stage behavioral deficits, including vocal communication and anxiety, that progress during mid-to-late adulthood (6–12 months of age). Yet, the biological pathways and mechanisms that lead to prodromal dysfunction are not well understood. This study investigated the Pink1−/− rat in young adulthood (2 months of age). Mixed sex groups of Pink1−/− rats and wildtype (WT) controls were assayed for limb motor, anxiety, and vocal motor behaviors. A customized NanoString CodeSet, based on genetic work in later adulthood, was used to probe for the up regulation of genes involved in disease pathways and inflammation within the brainstem and vocal fold muscle. In summary, the data show sex- and genotype-differences in limb motor, anxiety, and vocal motor behaviors. Specifically, female Pink1−/− rats demonstrate less anxiety-like behavior compared to male Pink1−/− rats and female rats show increased locomotor activity compared to male rats. Pink1−/− rats also demonstrate prodromal ultrasonic vocalization dysfunction across all acoustic parameters and sex differences were present for intensity (loudness) and peak frequency. These data demonstrate a difference in phenotype in the Pink1−/− model. Tuba1c transcript level was identified as a key marker negatively correlated to ultrasonic vocalization at 2 months of age. Identifying genes, such as Tuba1c, may help determine early predictors of PD pathology in the Pink1−/− rat and serve as targets for future drug therapy studies.

Objective: Radiation therapy (RT) for head and neck cancer (HNC) can result in severe xerostomia, or the subjective feeling of dry mouth. Characterizing xerostomia is critical to designing future clinical trials investigating how to improve HNC patients' quality of life (QoL). Few studies have investigated the very late (>5 years post‐RT) effects of RT for HNC. We undertook preliminary studies quantifying very late xerostomia.Methods: Six adults who underwent RT for HNC at least 5 years prior and reported xerostomia were enrolled. Five healthy adults without a self‐reported history of HNC or xerostomia were enrolled as controls. All participants completed three validated surveys to measure xerostomia‐related QoL. Salivary production rates were measured and compositional analysis of the saliva and oral microbiome was completed.Results: The QoL survey scores for the HNC participants were significantly worse as compared to the control participants. The HNC participants produced less unstimulated saliva (p = .02) but not less stimulated saliva. The median salivary mucin significantly higher in HNC participants than in control participants (p = .02). There was no significant difference between the pH, amylase, or total protein. Microbiome analysis revealed alpha diversity to be significantly lower in the HNC participants.Conclusion: In the survivors of HNC who suffer from late toxicities, multiple means of measuring toxicity may be useful. We found that in patients with radiation‐induced xerostomia over 5 years after therapy, not only were the QoL surveys significantly worse, as expected, but other measurements such as mucin and oral microbiome diversity were also significantly different.Level of evidence: 3.