Single-cell analyses of nitrergic neurons in simple nervous systems

Understanding the role of the gaseous messenger nitric oxide (NO) in the ne rvous system is complicated by the heterogeneity of its nerve cells; analys es carried out at the single cell level are therefore important, if not cri tical. Some invertebrate preparations, most especially those from the gastr opod molluscs, provide large, hardy and identified neurons that are useful both for the development of analytical methodologies and for cellular analy ses of NO metabolism and its actions. Recent modifications of capillary ele ctrophoresis (CE) allow the use of a small fraction of an individual neuron to perform direct, quantitative and simultaneous assays of the major metab olites of the NO-citrulline cycle and associated biochemical pathways. Thes e chemical species include the products of NO oxidation (NO2-/NO3-), L-argi nine, L-citrulline, L-ornithine, L-argininosuccinate, as well as selected N O synthase inhibitors and cofactors such as NADPH, biopterin, FMN and FAD, Diverse cotransmitters can also be identified in the same nitrergic neuron. The sensitivity of CE methods is in the femtomole to attomole range, depen ding on the species analysed and on the specific detector used. CE analysis can be combined with prior in vivo electrophysiological and pharmacologica l manipulations and measurements to yield multiple physiological and bioche mical values from single cells, The methodologies and instrumentation devel oped and tested using the convenient molluscan cell model can be adapted to the smaller and more delicate neurons of other invertebrates and chordates .