Elsevier

Behavioural Brain Research

Volume 301, 15 March 2016, Pages 226-237
Behavioural Brain Research

Research report
Investigation of diazepam efficacy on anxiety-like behavior in hemiparkinsonian rats

https://doi.org/10.1016/j.bbr.2015.12.045Get rights and content

Highlights

  • Hemiparkinsonian rats have higher anxiety-like behavior than non-parkinsonian rats.

  • Hemiparkinsonian rats have the greatest prevalence of ‘high anxiety’ behavior.

  • Diazepam is least efficacious in high anxiety hemiparkinsonian rats.

Abstract

There is growing recognition that anxiety disorders have a greater impact on quality of life in Parkinson’s disease than motor symptoms. Yet, little is known about the pathophysiology underlying this non-motor symptom in Parkinson’s disease which poses a considerable barrier in developing effective treatment strategies. Here, we administered diazepam to hemiparkinsonian and non-parkinsonian rats and assessed its efficacy in three anxiety behavioral tests. At present, no information about this exists in preclinical research with sparse data in the clinical literature. Moreover, diazepam is an acute anxiolytic which makes this drug a suitable research tool to unmask differences in anxiety-like behavior. Using the unilateral, medial forebrain bundle 6-hydroxydopamine rat model of Parkinson’s disease, we noted that hemiparkinsonian rats had more baseline anxiety-like behavior with 60% of them exhibiting high anxiety (HA) behavior in the elevated plus maze. In contrast, 41% of sham-lesioned rats and 8% of naïve rats exhibited HA behavior. Next, we employed the elevated plus maze and noted that diazepam (1.5 mg/kg) was anxiolytic in low anxiety (LA) sham-lesioned (p = 0.006) and HA sham-lesioned rats (p = 0.016). Interestingly, diazepam was anxiolytic for LA hemiparkinsonian rats (p = 0.017), but not for HA hemiparkinsonian rats (p = 0.174) despite both groups having similar motor impairment and parkinsonian phenotype. Overall, diazepam administration unmasked differences in anxiolytic efficacy between HA hemiparkinsonian rats, LA hemiparkinsonian rats and non-parkinsonian rats. Our data suggests that neuro-circuits involved in anxiety-like behavior may differ within these groups and posits that diazepam may have reduced efficacy in certain individuals with PD anxiety disorders.

Introduction

Clinically significant anxiety is a non-motor symptom in Parkinson’s disease (PD) that can affect up to 75% of PD patients [1], although most studies show a prevalence in the 20–40% range. This variability reflects the lack of a standardized PD-specific anxiety test [2], the wide spectrum of anxiety disorders in PD [3] and the relatively recent realization of this non motor-symptom of PD in routine clinical testing. Anxiety can burden the disease by significantly worsening parkinsonian motor symptoms [4] and negatively impacting quality of life [5], [6]. In fact, anxiety is also one of the best predictors of poor quality of life in PD and has been rated more harmful to well-being and more functionally detrimental to PD patients than the motor symptoms of this disease [7], [8].

While anxiety disorders in PD could be intuitively attributed to the diagnosis of a progressive and chronic neurodegenerative disease, it is still more prevalent in PD than in other chronic medical illnesses such as multiple sclerosis [1] and in the general population [9]. Several preclinical studies have also shown an increase in anxiety-like behavior in PD animal models where the psychological impact of living with a chronic disease is not a factor [10], [11], [12]. Additionally, while anxiety can occur at any stage of the disease [13], retrospective studies show that anxiety may serve as an early risk marker for PD since this non-motor symptom manifests during the prodromal stage, many years before the onset of motor symptoms and prior to the diagnosis of PD [10], [14], [15], [16], [17].

Since little attention has been given to understanding anxiety disorders in PD, there is a lack of awareness and knowledge about this non-motor symptom of PD and information from controlled trials for directing the best treatment option is lacking [18]. Therefore, it is not surprising that anxiety is under-diagnosed and untreated, with one study reporting 53% of PD patients remain un-medicated [19]. This has posed a critical barrier and challenge in developing effective treatment options for anxiety in PD and only a few studies have examined anxiety in preclinical rodent models of PD [10], [11], [12], [17], [20], [21], [22], [23], [24], [25]. Even fewer have tested whether typical anxiolytic medications, such as selective serotonin reuptake inhibitors, can diminish anxiety-like behavior with results showing little to no improvement [26], [27].

To date, the first-line treatment strategy for anxiety disorders specific to PD has not been established. Instead, current management strategies originate from personal observational studies and published guidelines based on the general public. Furthermore, there is no data to support any drug treatment for PD anxiety and the rationale is based on side effect profiles rather than efficacy [18], [28]. In this study, we sought to evaluate anxiety-like behavior in a 6-hydroxydopamine (6-OHDA) late-stage rat model of PD since it is uncommon for anxiety disorders to be diagnosed during the prodromal stage of the disease [13], and assess whether acute administration of diazepam would provide anxiolytic efficacy for two important reasons. First, diazepam is a benzodiazepine commonly prescribed as an anxiolytic for anxiety disorders in the general elderly population [29], but little data exists in clinical and preclinical literature regarding its efficacy in the acute treatment of PD anxiety. Second, the acute nature of diazepam efficacy provides a pragmatic approach in preclinical studies to reveal differences in anxiolytic responsiveness. Ultimately, findings from this study could reveal diazepam’s utility and/or limitation as a potential acute treatment option to alleviate anxiety in PD.

Section snippets

Rat model of PD

Animal use was conducted in accordance with the Albany Medical College Institutional Animal Care and Use Committee consistent with the National Institutes of Health guide for the care and use of Laboratory animals and all efforts were taken to minimize animal suffering and the number of animals employed in this study. Hemiparkinsonian rats were created by lesioning the right medial forebrain bundle as described previously [30]. Briefly, adult male Sprague Dawley rats (n = 164, 200–250 g, Taconic

TH immuno-reactivity and motor behavior

After injecting 6-OHDA into the right MFB, TH density in the right striatum and SNc was depleted by 96 ± 0.8% (n = 36) and 87 ± 2.4% (n = 33), respectively, compared to the intact (left) side (Fig. 1A–C, p < 0.0001, independent sample t-test). In the LAT, these rats exhibited forelimb akinesia by touching 89.4 ± 1.2% with the right un-impaired forepaw (n = 72 which included 6 hemiparkinsonian rats confirmed post hoc with striatal or SNc TH immuno-reactivity) whereas sham animals exhibited normal motor

Discussion

In this study, we found that hemiparkinsonian rats made fewer entries and spent less time in the open arm in the EPM than naïve or sham rats, implying that these animals have higher baseline anxiety-like behavior. These data suggest that 6-OHDA lesioning is anxiogenic. Further, when animals were subdivided into LA and HA groups based on baseline EPM behavior, we found that the vast majority of hemiparkinsonian rats could be classified as HA whereas the majority of naïve and sham rats could be

Conclusions

This study provides evidence that hemiparkinsonian rats have higher anxiety-like behavior with an increased prevalence of HA than naïve or sham rats. Furthermore, our novel findings show that HA hemiparkinsonian rats are less responsive to diazepam which may have important implications for anxiolytic treatment in PD patients. Future research should focus on aberrant neuronal signaling and pathophysiology in certain anxiety-related brain areas such as the amygdala and PFC to determine whether

Declaration of conflicting interests

Dr. Molho is a consultant for US World Meds and receives funding from Merz Pharmaceuticals, CHDI, Kyowa Hakko Kirin Pharma, US World Meds, Auspex Pharmaceuticals, Acadia Pharmaceuticals, Pfizer, and Civitas. Dr. Pilitsis is a consultant for Boston Scientific, Medtronic and St. Jude while receiving grant support from Boston Scientific, Medtronic, St. Jude and NIH 1R01CA166379. Dr. Ramirez-Zamora is a consultant for Medtronic and TEVA neuroscience and receives grant funding from Medtronic and

Acknowledgement

We would like to acknowledge and thank Dr. Lauren Jacobson for her expertise and guidance.

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