Stress is experienced throughout everyday life; when the body fails to reach homeostasis at a molecular, physiological, psychological and behavioural level in response to threatening stimuli. Recent experiments have shown that plant-derived essential oils possess anxiolytic and stress relieving properties in both humans and rodents. Two major components, cis-3-hexen-1-ol and trans-2-hexenal derived via biosynthesis from green leaves, have been found to produce physiological and behavioural responses consistent with a reduction in the body’s response to acute stress. Identification of these plant-derived compounds has lead to the development of Praescent™, a novel mixture of chemicals composed of cis-3-hexen-1-ol, trans-2-hexenal and α-pinene.
The main aims of the studies described in this thesis were to verify appropriate models of short and long-term stress in mice using physiological and behavioural measures and to investigate the effects of Praescent™ in alleviating the stress response. In addition, the effects of plant-derived odours were examined in a model of anosmic animals.
Rodents have commonly been restrained in single or repetitive sessions, which mimics the hormonal and behavioural stress response seen in humans. Therefore to address the first aim, chapter 3 investigated physiological (HR, cBT, locomotor activity and post-mortem studies on smooth muscle sensitivity to exogenous NAd) and behavioural measures in mice exposed to daily handling or restraint over a short or long-term period.
The results showed that simply handling mice over 21 days did not cause any disruptions in circadian rhythm, with higher levels of HR, cBT and locomotor activity during the active (dark) period. However at the onset of daily restraint, disruptions of normal circadian fluctuations of HR and locomotor activity were observed, with no changes in cBT. Alterations in the vas deferens responses to exogenous NAd were seen, with larger maximal contractions in tissues from stressed mice.
Behavioural measures showed that short-term, but not long-term, restraint stress induced anxiety-like effects, reflected by a reduction in the % number of entries and time spent in the open arms of the plus maze.
Furthermore, in chapter 3, an effective timing for treatment with Praescent™ was determined to develop an appropriate model for odour induced stress alleviation. Praescent™ induced anxiolytic effects when it was applied 3 days prior to stress and during the 3 days of stress procedure. No alleviation of stress was observed when the iii odour was applied on the last day of the 3 day stress, 30 min prior to measuring behaviour or throughout the 3 days of the stress period.
In chapter 4 the effects of plant-derived odours on a model of short-term stress were investigated. The results showed that short-term stress induced by daily restraint is anxiogenic and the behavioural changes which indicate anxiety are attenuated in mice which have been exposed to Praescent™, without affecting the level of locomotor activity. Exposure to the individual components of Praescent™ (cis-3-hexen-1-ol, trans-2-hexenal and α-pinene) also showed anxiolytic effects.
Chapter 5 examined the beneficial effects of plant-derived odours in a model of unpredictable long-term stress. Exposure to Praescent™ did not reverse the changes in circadian rhythm for HR. The post-mortem responsiveness of the vas deferens to NAd was similar to that in the control groups, suggesting that Praescent™ reduced the alterations in sympathetic nervous system, which occur in response to concurrent stresses.
The mechanisms of action by which the odours alleviate stress were examined in chapter 6. Mice were rendered anosmic by irrigation of zinc sulfate or by an intraperitoneal injection of methimazole, which resulted in degeneration of olfactory epithelium and failure to find a food pellet, buried in the bedding. However anosmic mice exposed to daily stress for 3 days did not demonstrate anxious behaviour. Therefore, it was not possible to determine whether the ability of the mice to detect the odour of Praescent™ was necessary for it to exert its anxiolytic effects.
In conclusion, the effects of laboratory procedures, such as restraint, on the physiology and behaviour of mice, were examined. Importantly, the therapeutic effects of plant-derived odours and their ability to palliate the negative effect of stress were demonstrated, indicating the potential use of Praescent™ as a non-sedative anxiolytic treatment to improve the welfare of experimental mice by reducing stress responses, and consequently improve the research data obtained from the animal models.