We will examine the vesicular traffic of lectins, toxins (Cholera toxin along with its exogenous GM1 ganglioside receptor and Ricin 60) and Nerve Growth Factor (NGF) into the Golgi apparatus of cultured neuroblastoma and rat pheochromocytoma cells (PC12), and in vivo in the rat nervous system. We propose that this vesicular transport pathway involving various ligands and, probably, their receptors mediates physiologic and pathologic reactions of the neuron. The endocytic pathway(s) in neurons will be studied in detail with quantitative autoradiographic and morphometric methods to investigate and contrast fluid-phase with adsorptive endocytosis and define the sequential transport of the ligand into the various endocytic compartments. Correlations between morphologic observations and biologic effects of the above ligands will be established with derivatized biologically active ligands suitable for morphologic studies. Experimental conditions will be modified to allow binding and endocytosis of the ligand but precluding expression of activity; thus we will be able to identify the compartment of the endocytic pathway which is crucial for the ligands biologic activity. The NGF induced morphologic modulation of the Golgi apparatus of PC12 cells will be explored further with morphologic methods and quantitative studies examining the number and affinities of plasma membrane receptors to NGF and Wheat germ agglutinin, and the magnitude of endocytosis of these ligands into the Golgi apparatus. Using antibodies against the Golgi apparatus the distribution of antigens in the Golgi apparatus of the neuronal perikaryon and along the smooth endoplasmic reticulum of axons and dendrites will be studied with light and electron microscopic, immunocytochemical, and autoradiographic techniques. The Golgi apparatus of rat cerebral neurons and NGF stimulated PC12 cells will be isolated for immunologic and enzymatic characterization. Existing anti-Golgi antibodies, produced by D. Louvard, reacting with a 135000 D antigen of the Golgi apparatus of several cell types will serve as controls for the isolation of neuronal Golgi apparatus and for the search of neuronal Golgi specific antibodies.
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