The proposed sabbatical research is a part of an overall program of research focussed on the identification and characterization of central and peripheral neural pathways responsible for control of food intake by glucose and fatty acids. Previous work strongly indicates that the neuropeptide, galanin, participates in the stimulation of feeding by decreased fatty acid oxidation. The specific goal of the proposed work is to further clarify the role of galanin in the nucleus of the solitary tract (NTS) in the stimulation of feeding by 2-mercaptoacetate (MA), a drug which antagonizes mitochondrial beta oxidation of fatty acids. The experimental approach will utilize Fos immunohistochemistry to label neurons in the NTS activated by MA and the retrograde tracer, cholera toxin conjugated to horseradish peroxidase (CT-HRP), to label vagal sensory terminals in the NTS. With these functional and anatomical labels, it will then be possible to determine whether vagal sensory neurons terminate on NTS cell bodies which express Fos in response to MA. An additional goal is to determine whether vagal sensory terminals making contact with Fos-immunoreactive cell bodies contain galanin, and if not, to determine which peptides they contain. In these studies, techniques already in use in the Flinders laboratory will be utilized for the combined application of CT-HRP with peptide and Fos immunohistochemistry at the light and electron microscopic level. These procedures employ the tetramethyl benzidine/ammonium paratungstate reaction, which is further stabilized with diaminobenzidine and cobalt chloride, to visualize the CT-HRP. For peptide immunochemistry, immunogold is employed as the reporter, as it forms a reaction product which is distinguishable from the CT-HRP and which is suitable for electron microscopy. The proposed experiments take advantage of the special anatomical expertise of Dr. Llewellyn-Smith and her colleagues (including Rds. Pilowsky and Minson) at Flinders University. This group has played a key role in the development of retrograde and anterograde tracers compatible with peptide immunohistochemistry for light and electron microscopic analysis. Their laboratory is world-renowned for their use and refinement of these and related techniques for analysis of central and peripheral pathways for control of blood pressure. Results of the proposed research will reveal anatomical and neurochemical organization underlying the control of food intake by fatty acid availability. This knowledge will lead to a fuller understanding of the relationship between appetite and metabolism and the neural substances which bridge these important physiological events.
