Ethological foundations of measuring obsessive-compulsive phenotypes in the deer mouse: a methodological perspective
De Brouwer, Geoffrey
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Pre-clinical models which leverage the study of animal behavior are invaluable tools to advance our understanding of a variety of psychological constructs. Such ethological investigations allow us to gain insight into abnormal biological processes which may contribute to the provocation of abnormal cognitive patterns which in turn may develop into a variety of ‘abnormal’ behaviors. Such behaviors, if allowed to proceed unchecked, typically progress into pathological states which interfere with the typical functioning of individuals, then termed psychiatric disorders. An example of such a disorder is obsessive-compulsive disorder (OCD), the focal point around which the investigations in this thesis were conceptualized. OCD is typically characterized by disruptions in both cognition (obsessions) and behavior (compulsions) leading to significant impairment in both executive function and quality of life. From an ethological perspective, OCD has been modelled in a handful of species applying several experimental frameworks and behavioral tests. Broadly speaking, these typically rely on the observation of repetitive behaviors and how they respond to established or prospective treatment strategies. As models become more established, investigations into the neurobiology of these modeled compulsive-like behaviors are initiated. Encouragingly, many analogous neurotransmission systems, receptors, brain regions and sub-circuits have been implicated in the expression of OCD-like behavior in these animal models, highlighting their translational relevance for the study of intricate psychiatric disorders such as OCD. However, a key short coming of such models remains the demonstration of the affective components of OCD, i.e. compulsions, within them. The species employed in the current investigation is the deer mouse (Peromyscus maniculatus bairdii), a validated but still rapidly developing model of OCD which offers select key advantages. Firstly, the compulsive-like behaviors exhibited by these animals are entirely naturalistic, not requiring any specific breeding, genetic knockout, pharmacological or behavioral induction. This exemplifies an ideal model, since it permits the careful study of abnormal behaviors in a context which is as similar as possible to the human condition, since the behaviors in question presumably originate naturally due to some unique underlying disturbance. Furthermore, deer mice naturally present with three different types of compulsive-like behaviors. These include excessive, highly stereotypical movements such as back flipping, the building of excessively large nests (LNB), and occurrence of high marble-burying (HMB). The two former behaviors have been validated by showing a response to chronic escitalopram (an SSRI; 50 mg/kg/day) treatment. As such, the current investigation set out to continue the development of the deer mouse model of OCD, by studying HMB, albeit indirectly, by dissecting the marble burying test (MBT), the behavioral test widely applied to test anti-compulsive and anti-anxiety drug action. This was done by means of an investigation into the methodological parameters of the test as well as a review of relevant literature pertaining to the application of the test. Secondly, LNB which represents a highly goal-directed behavior, ideal for the study of the motivational factors underlying compulsive murine behavior, was investigated in a novel experimental paradigm. The dissection of the MBT presented here represents a ‘back-to-basics’ approach concerning ethological studies. The test itself is relatively straight-forward, requiring a number of marbles to be placed on a flattened layer of bedding material (typically wood shavings) inside a test cage. Rodents are then introduced to these prepared cages and allowed a certain amount of time to interact. The number of marbles which are buried beneath the surface after the test session are tallied as an index of compulsive-like behavior. While being widely applied in the scientific literature, the MBT is striking in terms of the lack of methodological congruence between different laboratories. Resultantly, we argue here that special attention should be paid to even the most seemingly insignificant details of even well-established behavioral tests. Indeed, by investigating the use of several different burying substrates of differing densities in the MBT, we show that such a manipulation can robustly influence the outcomes of the test. Furthermore, we highlight that the subjective manner in which the test is scored is indeed subject to inter-observer variability. Following from the findings above, we performed a review of investigations in which the MBT was applied as the primary behavioral assay. Once again, it was highlighted that the test is employed with a wide range of experimental parameters in terms of the size of test cages, number of test repetitions, test duration, number and size of marbles, burying substrate and scoring criteria used. In terms of treatment response, the MBT reports anti-compulsive effects for a wide variety of drugs, some of which are not typically effective in the clinical setting. These findings are further complicated by the realization that many of these drugs, which require lengthy periods for onset of action, are administered acutely, often minutes before the test. Taking all of these findings into account, we argue for the standardization of many of the aforementioned variables, so as to improve the practical utility of the MBT as a screening test for anti-compulsive drug action. Another behavior exhibited by certain species such as mice and rabbits, concerns the excessive, diligent building of nests. As is the case with the MBT, the concept underlying LNB as a test for compulsive-like behavior is relatively simple. In brief, examining LNB involves the provision of an excess of nesting material to the housing cages of mice and recording the quantity of material that is used by each subject over the course of several nights. Animals that consistently use excessive quantities of material as appraised against the behavior of the larger population, are then selected for further investigation. Indeed, it has been shown that deer mice selected in this fashion show a reduction in nesting size following SSRI treatment. Since all mice that express LNB are housed under identical conditions to their normal counterparts, it stands to reason that there is some unique underlying dysfunction motivating the disinhibited building of nests. Towards investigating this premise, cohorts of normal and LNB were selected and tested in a novel testing environment. Here, the mice were placed into cages containing automated nesting material dispensers. To acquire nesting material, mice would have to execute lever-presses. After mice had learned this action-outcome association, their behavior was tested under two unique circumstances. Firstly, the nesting material was withdrawn from the dispensers so that lever-presses no longer resulted in any outcome. In the second instance, the levers delivered a mild electric shock each time a lever-press was executed. Interestingly, in this novel environment, LNB mice generally executed more lever presses than their NNB counterparts, particularly during the two unique phases described above. Mice were retested under identical circumstances following 28-days of chronic escitalopram treatment, showing preliminary evidence that the motivational drive to engage in nesting behavior abates, as was evinced by the lower number of lever-presses that were executed during the punishment phase. The study design above was intended to test whether LNB is characterized by unique cognition, inspired by the finding that OCD patients often show difficulty in changing their obsessive-compulsive behavioral routines, and that they are prone to endure negative consequences so that their compulsive rituals may be given attention. This represents a shift in ethological investigations, which in many cases rely primarily on appraising behavior at face-value, without attempting to divulge the cognitive factors which may play a role in the presence of such behaviors which are considered abnormal in the first place. In conclusion, this thesis argues that ethological studies should be continually refined to improve the validity of the findings made, regardless of how simple the core premise of the given model or behavioral test may be. Furthermore, the foundational basis of ethological studies should be conceptualized bearing in mind that animal behavior is often more conscientious than many behavioral tests take into account, and that careful study design can leverage this to carefully investigate behaviors at a deeper level, which in turn can add to our knowledge of psychiatric disorders.
- Health Sciences