Pharmacological evaluation of an alpha2C selective antagonist in an animal model of posttraumatic stress disorder

Abstract

Posttraumatic stress disorder (PTSD) is a psychiatric disorder that can manifest following the experience or witnessing of a life-threatening event such as military combat, natural disasters, terrorist incidents, serious accidents, or physical or sexual assault during child- or adulthood. Although most survivors of a traumatic event recover over time, approximately 30% of victims will go on to develop PTSD. Dysfunctional activity of multiple neurobiological pathways has been implicated in the pathology underlying PTSD symptomatology, including the noradrenergic, serotonergic, dopaminergic and glutamatergic systems, as well as theHypothalamic-pituitary-adrenal (HPA) axis. These systems are also mutually interlinked, with dysfunction of one system affecting the function of the other, thereby complicating the pathology of the disorder. This complexity demands deeper investigation to define the roles of each system in the neuropathology of PTSD. The noradrenergic system is prominent and represents an important pharmacological target in attempts at preventing the development of PTSD in the immediate aftermath of trauma. In PTSD, it has been found that emotional events are associated with high levels of noradrenalin (NA) release in brain areas involved in learning and memory such as the amygdala and hippocampus. Emotional memories are mainly influenced by noradrenergic α1/2 and β2 receptors. In the basolateral amygdala, emotionally aroused noradrenergic activation tends to strengthen memory consolidation in the hippocampus which is responsible for arranging contextual fear memory. Therefore, suitable curbing of the noradrenergic system could be an important neurobiological target in treating PTSD. The first objective of this study was to validate the predator scent exposure (PSE) model in our laboratory and to determine if male Wistar rats will demonstrate different levels of anxiety akin to maladaptation and well-adaptation following exposure to a traumatic event, i.e. predator scent. In this regard, we identified the extreme ends of the normal distribution of anxious behaviour in response to PSE to validate the PSE model, as described below. That said, the number of faecal boli passed during PSE was used as a measure of immediate trauma-induced anxiety. To this end, the elevated plus maze (EPM) have been used to establish the application of the PSE model as a valid framework for novel drug discovery. Parameters assessed in the EPM included number of entries into the open (OAE) and closed (CAE) arms, as well as the time spent in the open (sOA) and closed (sCA). Head-dipping behaviour in the open arms was applied as a measure of risk assessment and exploratory behaviour. Head dipping occurs in the central area of the maze and reflects an aversion to risk taking, which is related to the anxious state of the animal. The PSE model has ethological relevance as it mimics intense stressful experiences and results in long term changes in behavioural, autonomic and hormonal responses that correlate with the symptoms in human PTSD. Regarding the conceptual validity of the model, "predator exposure trauma" is a potentially life-threatening situation and may represent a more "natural" challenge than other forms of stressors, i.e. electric tail shocks or restraint that may in fact be more related to extreme conditions such as torture. To confer validity to the model, male Wistar rats were used because of their recognised enhanced sensitivity to stress. Wistars were exposed individually under dim white light conditions (15 lux) for 10 minutes to a 10 cm x 10 cm cloth previously exposed to a male cat for 2 months, and a control group that were exposed to a clean non-cat-scented 10 cm x 10 cm cloth, with both groups being tracked and recorded digitally for subsequent analysis using Ethovision XT® software. Evidence from this study confirmed that at least 20 — 25% of the exposed group develop anxiety-like behaviours in the EPM when exposed to cat scent, while the remainder of the group was regarded to be resilient. This finding is important since individual variation in and susceptibility to trauma-related pathology is a key criterion for an animal model of PTSD. This effectively validated the animal model on face value in our laboratory for further application in this study. Given the causal role for the hypothalamic-pituitary-adrenal (HPA) axis in PTSD, the second objective of this study was to compare plasmacorticosterone concentrations between treatment-naive well-adapted and maladapted animals as assessed 21 days post PSE. Moreover, we also sought to determine whether the selective α2C AR antagonist, ORM-10921, was able to modify plasma corticosterone in PSE animals when dosed immediately post-PSE for 21 days. ORM-10921 treatment induced a subtle albeit insignificant trend towards hypocorticortisolemia however upon close scrutiny ORM-10921 treated animals were characterised by an increase in plasma corticosterone and an associated lowering of anxiety compared to vehicle treated cohorts. Finally, considering the prominent role for NA in the neurobiology and treatment of PTSD, the third objective of this study was to investigate whether the selective α2C AR antagonist, ORM-10921, is capable of reversing PSE-induced anxiety-like behaviour related to PTSD. Considering the less than adequate response of clinical PTSD to modulation of the noradrenergic and serotonergic systems, i.e. chronic treatment with inter alia TCAs, SNRIs, and SSRIs, as well as disappointing findings regarding the prophylactic use of propranolol in PTSD, there is increasing need for novel drug treatments that will offer improved and sustained efficacy. The current investigation found that administration of 0.3 mg/kg ORM-10921 only from 8 days after exposure (Addendum C) effectively increased behavioural disruptions evident in the PSE model, as evinced by a decrease in number open arm entries (OAE) and a decrease the time spent in open arms (sOA); this was also found in saline treated groups. Interestingly, ORM-10921 dosed in this manner tended to elevate corticosterone levels, which coincides with some evidence in the EPM that delayed-onset ORM-10921 administration is anxiogenic. That said, administration of a 0.3 mg/kg immediately after exposure (Chapter 3) reduced behavioural disruptions that model PTSD, with chronic ORM-10921 treatment significantly lowering anxiety like-behaviours evinced by increases in the number of open arm entries(OAE) and time spent in open arms (sOA) compared to the saline treated animals. ORM-10921 also increased risk assessment and exploratory behaviour compared to saline treated animals as evident in the number of head dipping episodes. Concluding, this study has conferred construct, face and perdicitve validity to the PSE model established in our laboratory and confirms the model's status as a prominent animal model in PTSD. Moreover, it has provided greater insight into the role of noradrenergic receptors in anxiety related PTSD by providing supportive evidence that selectively blocking the α2C AR inhibits PTSD-related behaviour, notably so when administered immediately post-trauma. ORM-01921 therefore may be a viable secondary treatment option to prevent the development of PTSD post trauma. Further studies employing these novel agents in the treatment of anxiety disorders, such as PTSD, are encouraged, and will further our understanding of the role of α2C AR in such disorders, as its viability as a therapeutic target.