Characterization of chemically defined neurons and their cellular relationships by combined immunocytochemistry and radioautographic localization of transmitter uptake sites

Abstract

Radioautography and immunocytochemistry may be combined at the light and electron microscopic levels for simultaneously localizing uptake sites for exogenous transmitter molecules [such as (3H)monoamines or (3H)amino acids] and endogenous transmitter-related antigens (classical transmitters and their synthesizing enzymes as well as neuropeptides) in the central nervous system. Silver grain accumulations indicative of transmitter uptake sites are readily distinguishable from immunocytochemical labels of the peroxidase-antiperoxidase (PAP), avitin-biotin, or colloidal gold methods. The combination of uptake radioautography and immunocytochemistry may be applied to the investigation of (1) the chemical identity of (3H) transmitter-accumulating elements, (2) the coexistence of different neurotransmitters within the same neurons, and (3) the cellular basis of interactions between certain neurotransmitters, in particular monoamines, GABA, and neuropeptides. This article describes and evaluates the method and reviews the available experimental data derived from its application.

Selective Labeling of Visual Corpus Callosum Connections with Aspartate in Cat and Rat?

“…In the present study, ipsilateral labeling occurred after 24 and 48-72 h survivals in rat and cat, respectively; thus, the absence of contralateral GABA label was not due to a failure of GABA transport. Some investigators suggest that studying GABA neurons is best done with a pretreatment of amino-oxyacetic acid (AOAA) to prevent intraneuronal GABA breakdown (Harandi et al, 1986;Iversen & Schon, 1973) but this method has been questioned by other investigators (Bosler et al, 1986). Even when investigators have compared GABA labeling in animals with and without AOAA treatment, no differences are found (Kisvarday et al, 1986;Levi et al, 1983), therefore the absence of AOAA pretreatment in the animals of the present study does not account for the negative result in terms of contralateral labeling.…”

Neuronal imaging with colloidal gold

“…Similarly, postembedding immunogold could be used to identify the putative neurotransmitter of neurons whose cell type had been characterized by Golgi impregnations . To study the morphological relationship between neurons that have high uptake mechanisms for neurotransmitters and those that contain immunocytochemically identifiable neurotransmitters, ultrastructural autoradiography can be combined with postembedding colloidal gold immunostaining (Bosler et al, 1986). Immunogold staining can also be combined with peroxidase immunolabelling to facilitate light microscopic analysis of neuritic trees after plastic embedding but prior to selection for ultrastructural analysis (see Fig.…”