An investigation of the effects of fentanyl on respiratory control

Kennedy, Ashleigh (2015) An investigation of the effects of fentanyl on respiratory control. PhD thesis, University of Glasgow.

Abstract

Respiration is a complex rhythmic motor behaviour that metabolically supports all physiological processes in the body and is continuous throughout the life of mammals. A failure to generate a respiratory rhythm can be fatal. Understanding how the respiratory rhythm is generated by the brainstem presents a substantial challenge within the field of respiratory neurobiology. Studies utilising in vitro and in vivo rodent models have provided compelling evidence that a small bilateral region of the ventrolateral medulla, known as the preBötzinger complex (preBötC), is the site for respiratory rhythmogenesis. There is also evidence to suggest a second distinct neuronal group, the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG), plays a specialised role in respiratory rhythm generation in the neonatal rodent. During early life in rodents and humans, the respiratory system is immature and an irregular breathing pattern is generated, making this period of life potentially vulnerable to external perturbations. However a step in maturity occurs early in life after which breathing becomes regular. Currently, the underlying mechanisms involved in respiratory rhythm generation during early life are not fully understood. It is hypothesised that the RTN/pFRG functions as the dominant respiratory rhythm generating oscillator during early life when the respiratory system is immature, after which the preBötC becomes the dominant rhythm generator. However, how the preBötC and the RTN/pFRG interact in vivo to produce rhythmic breathing during postnatal development remains elusive.

The first aim of this thesis was to assess postnatal maturation of breathing patterns in the mouse using non-invasive whole body plethysmography. Between postnatal day (P) 2 and P3, a critical maturation step occurred, whereby breathing transitioned from an unstable and dysrhythmic pattern to a regular and robust pattern. The second aim of the thesis was to investigate the influence of this postnatal maturation on central respiratory control. Mu (μ) opioid receptor agonists are known respiratory depressants. The activity of the preBötC is depressed by μ opioids in vitro. Furthermore, fentanyl, a potent μ opioid receptor agonist, evokes respiratory frequency depression in vivo by exclusively targeting and depressing preBötC neurons. Conversely, the RTN/pFRG is insensitive to μ opioids. Accordingly, fentanyl was utilised as a pharmacological tool to selectively perturb the preBötC in vivo throughout postnatal development and through to early adulthood. The acute respiratory depressive effects of fentanyl were measured in order to investigate the level of involvement of the preBötC in respiratory rhythm generation throughout this critical developmental time period. Based on the general hypothesis that the preBötC functions as the dominant respiratory rhythm generator when the respiratory system has matured, it was hypothesised that mice would be more susceptible to the respiratory depressive effects of fentanyl after the maturation step has occurred i.e. the respiratory sensitivity to fentanyl would be age-dependent. Initially, mice were repeatedly exposed to fentanyl throughout postnatal development. However, fentanyl failed to induce a respiratory depression at all postnatal ages, suggesting repeated exposure had induced a rapid desensitisation to fentanyl’s respiratory effects. The study design was consequently altered to allow the hypothesis to be sufficiently tested, whereby different mice were studied on each postnatal day i.e. each mouse was only exposed to fentanyl once. This study revealed a trend towards an age-dependent increase in respiratory sensitivity to fentanyl, where mice displayed a heightened respiratory frequency depression in response to fentanyl after the maturation step had occurred from P3 onwards. This data therefore lends support to the hypothesis that the preBötC functions as the dominant respiratory rhythm generator post-maturation.

In the clinical setting fentanyl is widely utilised for treating chronic and acute pain. However, despite the potent respiratory depressive actions of fentanyl, the long-term respiratory consequences of repeated exposure remain unexplored both clinically and pre-clinically. Owing to the immaturity of the respiratory system and the corresponding fragile nature of breathing patterns during neonatal life in mammals, a further aim of the thesis was to determine the long-term effects of fentanyl exposure during this vulnerable respiratory time period in the mouse. To establish if the postnatal age of fentanyl-exposure influences long-term respiratory effects, fentanyl exposure during juvenile life, which is regarded as being post-respiratory maturation, was also assessed. Neonatal mice were exposed to fentanyl (0.04 mg/kg daily) from P1-P5 and juvenile mice were exposed from P9-P13. When mice reached adulthood, baseline respiratory activity and the respiratory response to a subsequent fentanyl challenge were assessed during wakefulness and under anaesthesia. When awake, neonatal-exposed mice exhibited a reduced baseline respiratory frequency and an attenuated respiratory sensitivity to fentanyl. Under anaesthesia, neonatal-exposed mice displayed a depressed baseline minute ventilation and a high frequency of spontaneous augmented breaths. In direct contrast to the wakeful state, when anaesthetised, neonatal-exposed mice exhibited a striking hypersensitivity to the acute respiratory depressive actions of fentanyl. In all neonatal-exposed mice, fentanyl evoked a respiratory failure. In juvenile-exposed mice, baseline respiratory activity remained unaltered in the wakeful state and fentanyl also failed to induce a respiratory depression. When anaesthetised, baseline minute ventilation remained unchanged and the high occurrence of augmented breaths exhibited by the neonatal-exposed mice was not observed. Unlike the wakeful state, fentanyl evoked a depression of respiratory activity in the juvenile-exposed mice when anaesthetised, however the augmented sensitivity to fentanyl and consequential respiratory arrest displayed by the neonatal-exposed was not observed. This data indicates that the anaesthetised state is more susceptible to respiratory depression. Furthermore, the data suggests that neonatal life represents a time period that is particularly vulnerable to the respiratory effects of opioid depression. The final aim of the thesis was to determine the long-term effects of neonatal fentanyl exposure on neurokinin-1 (NK1R) and μ opioid receptor expression within the ventral respiratory column (VRC), a region of the ventrolateral medulla comprising the preBötC. Neonatal-exposed mice exhibited significantly less NK1R and μ opioid receptor expressing cells in the region of the preBötC. This data suggests that repeated fentanyl exposure in neonatal life induces a long-term downregulation of these receptors.

In conclusion, fentanyl’s acute respiratory effects were age-dependent, which lends supports to the hypothesis that the preBötC functions as the dominant rhythm generator post-maturation. Furthermore, this thesis highlights the vulnerabilities of neonatal life to the lasting effects of opioid respiratory depression, whilst also providing invaluable insight into state-dependent respiratory modulation and depression.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Respiratory control, fentanyl, breathing, preBotzinger complex, RTN/pFRG, respiratory depression, in vivo, mouse, plethysmography
• Subjects: Q Science > QP Physiology
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Psychology & Neuroscience
• Supervisor's Name: McKay, Dr. Leanne
• Date of Award: 2015
• Depositing User: Miss Ashleigh Kennedy
• Unique ID: glathesis:2015-5998
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 10 Feb 2015 11:54
• Last Modified: 19 Feb 2015 10:52
• URI: https://theses.gla.ac.uk/id/eprint/5998