Neurotransmitter histochemistry: comparison of fluorescence and immunohistochemical methods

The above procedures clearly demonstrate the effectiveness of the fluorescence histochemical and immunohistochemical methods in analysis of central CA neuron systems. Each method is well characterized and possesses sufficient versatility to permit a variety of experimental applications. The fluorescence histochemical technique is extremely reliable, produces good cell and fiber morphology, and has served as the fundamental procedure used to define the organization and distribution of CA-containing neurons throughout both the central and peripheral nervous system. However, the basic method makes no distinction between individual catecholamine neuron systems unless combined with mechanical or chemically induced lesions. The immunohistochemical technique relies upon the availability of antisera generated against the catecholamine-synthesizing enzymes tyrosine hydroxylase (TH), dopamine beta-hydroxylase(DBH), and phenylethanolamine N-methyltransferase (PNMT). Each of these enzymes catalyzes different steps in catecholamine metabolism and therefore can be used in conjunction with one another to selectively delineate the organization and distribution of neuronal cells and processes containing dopamine (TH), noradrenaline (TH and DBH), and adrenaline (PNMT). In addition, the immunohistochemical method may be coupled with the fluorescence histochemical technique, dual-labeling immunohistochemical methods, and retrograde tract tracing methods to provide further information on the interrelations of individual CA systems with one another and other chemically distinct systems of neurons. The usefulness of the immunohistochemical method in elucidating the organization of separate systems of CA-containing neurons is illustrated in a recent study by Swanson and collaborators. Utilizing antisera raised against the previously mentioned CA synthesizing enzymes, they analyzed the organization of CA systems within the paraventricular and supraoptic nuclei of the hypothalamus. By combining this analysis with immunohistochemical staining with antisera generated against vasopressin and oxytocin, they were able to demonstrate differential distribution of adrenergic and noradrenergic fibers within each nucleus which could be correlated with the distribution of vasopressin-containing neurons. In addition, through the combined use of immunofluorescence and retrograde transport of the tracer dye true blue, they were able to show that a small percentage of TH-containing neurons within the paraventricular nucleus project to the region of the dorsal motor vagal complex and/or thoracic levels of the spinal cord. Although the later finding relied upon fluorescence microscopy, Bowker a