Mechanisms of mechanotransduction by dorsal root ganglia neurons
The molecular mechanisms that mediate mammalian sensory mechanotransduction are poorly understood. Detection of mechanical events by sensory neurons of the dorsal root ganglia (DRG) is the primary event in the senses of touch, pressure-induced pain and proprioception. Recent work has demonstrated that the somatic membrane of cultured DRG neurons is a suitable system for studying physical transduction. In this thesis the responses of cultured DRG neurons to focal mechanical stimulation were investigated. It was shown that mechanical stimulation activated non-selective cation channels in these cells. The response properties of different subclasses of sensory neurons were characterised and were consistent with the presumed in vivo phenotypes of these cells. A number of antagonists of mechanically activated currents, with affnity in the low micromolar range, were identified; these included the pore blocking compounds gadolinium and ruthenium red and FMl-43 acted as a permeant blocker of mechano- sensitive channels. Modulation of mechanically activated currents by extracellular calcium was observed and it was shown that currents were regulated by the actin cytoskeleton and the extracellular matrix protein laminin. Investigation of null mutant mice revealed that the acid sensing ion channels 2 and 3, which are widely hypothesised to function in mammalian mechanosensation, did not contribute to mechanically activated currents. Venom of the marine snail Conus ventricosus was found to block mechanically activated currents although the active component of this venom is yet to be identified. Overall, this work has shown that cultured DRG neurons are a useful system for studying mechanotransduction and has revealed a number of functional and pharmacological properties of the ion channels that underlie this process.