Mu-opioid receptor desensitization in ventral tegmental neurones: cellular mechanisms and functional consequences

Opiate drugs such as morphine and heroin are widely abused, with approximately 270,000 problem opiate users in the UK. When somebody uses opiates consistently for a while, they become tolerant to its effects and need to use more to get the ?high? they initially experienced and to stave off withdrawal. This process of tolerance contributes to the health and social problems inherent in opiate addiction. To date the mechanism(s) underlying the development of morphine tolerance remain unknown. In this proposal we will investigate opiate tolerance on a single cellular level, in order to investigate potential intracellular signalling mechanisms that may underlie the development of opiate tolerance. We will then go on to study how interfering with these signalling mechanisms can affect the development of opiate tolerance. It is only by understanding how morphine tolerance occurs on a cellular level that we can begin to design better treatments for opiate abuse in the future.

Opiate drugs are widely abused for their euphoric properties. Repeated administration of opiate drugs such as morphine or heroin results in the development of tolerance whereby the effects of a given dose are decreased, and therefore to obtain the same level of response a greater dose must be administered. To date the mechanism(s) underlying the development of opiate tolerance remain poorly understood. Opiates exert their effects by activating the G-protein-coupled mu-opioid receptor (MOR) in the CNS. MORs in common with many other G-protein-coupled receptors (GPCRs) can undergo a process of desensitization, leading to the speculation that this might be the mechanism underlying opiate tolerance.

Although the development of addiction to drugs of abuse is complex and involves many brain regions and processes, the initial euphoric, rewarding properties of opiates are closely linked to activation of MORs in the ventral tegmental area (VTA). MORs in the VTA are present on GABAergic neurones both somatodendritically (ie. on cell bodies and dendrites) as well as on nerve terminals. Activation of these receptors in both loci act to disinhibit dopaminergic neurones that innervate the nucleus accumbens and forebrain regions, activation of these pathways playing a key role in the development of reward and positive reinforcement. To date there have been no studies designed to investigate MOR desensitization in GABAergic VTA neurones. Crucially, there has been no definitive study investigating desensitization of GPCRs in general, located on nerve terminals of mammalian central neurones. This study will first investigate the molecular mechanisms underlying agonist-induced MOR desensitization in GABAergic VTA neurones using brain slice electrophysiology and viral-mediated gene transfer. We will determine the intracellular mechanism(s) underlying MOR desensitization in the VTA caused by morphine and DAMGO. Having determined the molecular mechanism(s) underlying agonist-induced MOR desensitization in the VTA, we will then go on to establish the functional consequences of desensitization of these receptors by investigating tolerance to the rewarding effects of opiates using conditioned place preference (CPP). Opiate CPP will be induced both under control conditions, and when MOR desensitization in the VTA is inhibited, either pharmacologically or using viral-mediated gene transfer.

Using this combination of electrophysiological and behavioural techniques, we will determine the molecular mechanisms underlying tolerance to the euphoric and rewarding properties of opiates. Only with a detailed knowledge of how tolerance occurs can we begin to design drugs that will either reduce or enhance the level of morphine tolerance.