Cytochemistry of undamaged neurons transporting exogenous protein in vivo
Authors: Broadwell Richard D. , Brightman Milton W. 
Published: January 2005

The uptake, transport and fate of exogenous protein in undamaged cranial motor and neurosecretory neurons in mice have been investigated using enzyme cytochemical techniques for horseradish peroxidase (HRP) and lysosomal acid hydrolases. Labeling of these neuronal perikarya with HRP is accomplished by retrograde axoplasmic transport from their axon terminals following vascular injection of the protein or by direct somal‐dendritic uptake subsequent to cerebral ventriculo‐cisternal perfusion of peroxidase. Axon terminals of neurosecretory neurons innervating the posterior pituitary gland and those of cranial motoneurons can remain unexposed to HRP administered intraventricularly and, therefore, cannot directly incorporate the protein.

Perikaryal organelles containing HRP reaction product include multivesicular bodies, small vacuoles, membrane‐delimited cisterns and dense bodies. The concentrations of these different types of peroxidase‐positive organelles are consistently greater in retrogradely labeled cell bodies than in perikarya which have pinocytosed the protein from the perisomal clefts, even though the amount of HRP bathing the cell bodies is much greater than the amount exposed to the axon terminals. Since the same groups of dense bodies contain lysosomal acid hydrolases as well as exogenous peroxidase, the HRP‐labeled dense bodies are indeed lysosomes. Exogenous peroxidase does not stimulate lysosome proliferation; the concentration of acid hydrolase‐positive lysosomes appears to remain the same whether or not the cells are labeled with HRP.

Despite peroxidase labeling of cranial motor and neurosecretory cell bodies after intraventricular injection, the anterograde axonal transport of peroxidase in these neurons is negligible. This transport can be demonstrably increased in the neurosecretory system, but not in cranial motor nerves, in response to hyperosmolarity induced in the animals by salt loading. Under such a stimulus many axons and Herring bodies in the posterior pituitary gland contain HRP‐labeled cisterns and 1,200–2,000 Å wide dense bodies. Both types of organelles also contain acid hydrolase activity and are confluent with each other. When compared to normal controls, the concentration of acid hydrolase‐positive organelles in pituitary stalk axons and Herring bodies from osmotically stressed mice is noticeably increased. Axonal cisterns with acid hydrolase activity and those transporting peroxidase in an anterograde or a retrograde direction appear similar morphologically.

Our results indicate that peroxidase pinocytosed by the neuron is eventually sequestered within perikaryal lysosomes for enzymatic degradation. Under normal conditions acid hydrolase activity and the majority of lysosomes are confined largely to dendrites and the cell body, and anterograde axonal transport of HRP is minor. In the osmotically stressed neurosecretory system, HRP appears to be carried coincidentally and anterogradely along with the movement of acid hydrolases, most likely from secondary lysosomes in the cell body. The movement of acid hydrolases down the axon is presumably increased for degradative purposes in the posterior pituitary gland. A part of the cisternal network conveying acid hydrolases and transporting peroxidase in anterograde and retrograde directions may represent a special compartment of agranular reticulum specifically involved with the lysosomal system of organelles and important in the segregation of exogenous macromolecules within the neuron for transport to or from lysosomes.