Rats that had one eye removed on the day of birth were examined at various postnatal ages with immunocytochemical methods to determine the effect on the development of the GABAergic axonal plexus in the visual cortex. The monocular segment of visual cortex contralateral to the enucleated orbit had 20-30% fewer GABAergic axon terminals than the monocular segment of visual cortex contralateral to the normal eye. Other cortical areas did not show any significant changes. These findings suggest that sensory deprivation of the visual cortex interferes with the normal development of GABAergic neurons.

Acetylcholinesterase (AChE) histochemical staining occurred in neurons of the dentate gyrus at the day of birth and steadily increased in intensity and distribution during the first 3 postnatal weeks until the adult pattern was reached. Granule cells failed to display AChE staining; however, the somata of most non-principal cells in these regions showed AChE activity. It is interesting that most hilar neurons in the dentate gyrus were AChE-positive, but molecular layer local circuit neurons and pyramidal basket cells associated with the granule cell layer did not display AChE staining. AChE reaction product was localized to the nuclear envelope and cisternae of the granular endoplasmic reticulum in the labeled neuronal somata. In addition, the neuropil in the dentate gyrus displayed AChE staining associated with membranes. The possible cholinoceptive role of the AChE somata in the hilus is discussed. © 1987.

Distributions of somata and neurites of cholinergic neurons were studied after seeding dissociated cells onto organotypic slice cultures. Slice cultures were made from hippocampal formation and adjacent cortical regions from rats or mice. Dissociated cell suspensions of basal forebrain tissue from rat or mouse fetuses were seeded onto the slice cultures. Combined cultures were maintained for 1-21 days in vitro. Cultures processed for acetylcholinesterase (AChE) histochemistry demonstrated non-random patterns of cholinergic cells and their neurites. Labeled cells appeared most frequently in the molecular layer of the dentate gyrus, and in the deeper layers of cortical regions adjacent to the hippocampus. Neurites extending from these labeled cells appeared to target the dentate molecular layer and the cortical subplate layer. By 4 days in vitro, AChE-positive basal forebrain cells display several short and thick neurites that appear to be dendrites, and one long process that appears to be an axon. By 5 days in vitro, dendrites are well developed; by 7 days the presumed axon has extended widely over the cortical target zone. These neurites are maintained through 3 weeks in culture. Distributions of cells varied with the age of the slice. AChE-labeled cells were not seen overlying hippocampal tissue when dissociated cells were seeded on slice cultures made from day 0 rats, but a few labeled cells were seen when seeded on slices from day 2 rats. Clear non-random patterns of labeled cells and neurite outgrowth were seen on slice cultures from day 5 or older pups. The non-random distribution seen with AChE-positive neurons was not seen using other techniques that labeled all cells (non-selective fluorescent labels) or all neurons; these techniques resulted in labeled cells scattered apparently homogenously across the slice culture.These studies demonstrate a non-random pattern of attachment or differentiation of basal forebrain cholinergic neurons when these cells are seeded onto cultured cortical slices; this pattern mimics the normal patterns of basal forebrain cholinergic projections to these cortical regions. These data suggest that the factors that normally guide basal forebrain-derived cholinergic axons to their target cells in vivo are present and detectable in this model system.

Intravascular injections of fluorescent or biotinylated tomato lectin were tested to study labeling of vascular elements in laboratory mice. Injections of Lycopersicon esculentum agglutinin (tomato lectin) (50–100 µg/100 µl) were made intravascularly, through the tail vein, through a cannula implanted in the jugular vein, or directly into the left ventricle of the heart. Tissues cut for thin 10- to 12-µm cryostat sections, or thick 50- to 100-µm vibratome sections, were examined using fluorescence microscopy. Tissue labeled by biotinylated lectin was examined by bright field microscopy or electron microscopy after tissue processing for biotin. Intravascular injections of tomato lectin led to labeling of vascular structures in a variety of tissues, including brain, kidney, liver, intestine, spleen, skin, skeletal and cardiac muscle, and experimental tumors. Analyses of fluorescence in serum indicated the lectin was cleared from circulating blood within 2 min. Capillary labeling was apparent in tissues collected from animals within 1 min of intravascular injections, remained robust for about 1 h, and then declined markedly until difficult to detect 12 h after injection. Light microscopic images suggest the lectin bound to the endothelial cells that form capillaries and endothelial cells that line some larger vessels. Electron microscopic studies confirmed the labeling of luminal surfaces of endothelial cells. Vascular labeling by tomato lectin is compatible with a variety of other morphological labeling techniques, including histochemistry and immunocytochemistry, and thus appears to be a sensitive and useful method to reveal vascular patterns in relationship to other aspects of parenchymal development, structure, and function.