Research objectives and content
The project will investigate the cellular and molecular bases of signal transduction in primary pain-sensitive neurones (nociceptors). Sensory transduction in nociceptors is poorly understood by contrast with transduction in other sensory receptors. The project will extend our understanding of a sensory system which is in many ways different from others. A better understanding of the mechanisms involved in the encoding of painful stimuli will also be of benefit in the development of improved analgesics and in the treatment of chronic pain.
The principal focus will be on the intracellular signalling pathways in pain-sensitive neurones which underlie the responses to the following pain-causing (i.e. algogenic) stimuli; (i) the peptide bradykinin, which is known to be released after tissue damage. Bradykinin serves the dual function of both causing pain and of enhancing the pain caused by other stimuli. (ii) noxious heat, which has been shown by the host group to activate an inward membrane current in nociceptors. The inward current is enhanced by bradykinin in a process which involves activation of protein kinase C.
The project will use sensory neurones isolated and maintained in culture. These neurones have been shown by the host laboratory and others to duplicate in many respects the properties of pain-sensitive neurones in intact animals, and they offer a convenient and powerful method for studying the cellular and molecular bases of the pathways responsible for transducing a painful stimulus into electrical excitation of the pain-sensitive neurone and subsequently into a sensation of pain. Responses of neurones to the stimuli outlined above will be dissected using electrophysiological techniques (whole-cell patch clamp) calcium imaging using confocal microscopy, and the methods of molecular biology (transgenic animals with knockout of specific genes such as the B2 receptor and PKC isoforms which may be important in pain transduction. production of tagged proteins used to follow transduction events such as protein kinase C translocation).
Training content (objective, benefit and expected impact)
The objective is a complete understanding of an important and surprisingly poorly investigated area of sensory transduction. The applicant would like to underline the practical importance of the possible results, in particular from a therapeutic point of view. The expected benefit to the applicant, already expert in electrophysiology, is the training in new and competitive
techniques deriving from the multidisciplinary approach with which this project will be carried out. that is PCR, confocal imaging,
immunohistochemistry, in situ hybridization, use of transgenic knockout animals. cell culture etc. The applicant is expecting to take back to his home lab in Italy the knowhow deriving from the above project. Links wth industry / industrial relevance (22)
An enhanced understanding of the pathways underlying the generation of pain has tremendous potential importance for the development of analgesics and anaesthetics by the drug industry. The host group and the applicant are in touch with Merck Inc. with a view to obtaining research funding for this work in view of their interest in bradykinin receptors. Knockout mice for the B2 bradykinin receptor subtype have already been supplied from Merck to the host group and will be a useful tool for the applicant.