Cytochrome oxidase staining marks dendritic zones of the rat olfactory bulb external plexiform layer

Cytochrome oxidase staining of the rat olfactory bulb external plexiform layer (EPL) produces a darkly stained intermediate zone bordered by lightly stained superficial and deep zones. Similar zonal staining was seen in cats, rabbits, and hamsters. These zones vary in relative thickness around the circumference of the olfactory bulb; the deep zone is proportionally thicker in the most dorsal and ventral parts of the bulb. Tufted cell somata are unevenly distributed within the EPL; the outer part of the EPL has more somata. The distribution of the cytochrome oxidase reaction product shows that the darkly stained intermediate zone is not produced by staining of tufted cell somata. Zones of cytochrome oxidase staining correspond to the sublaminar distribution of mitral and tufted cell basal dendrites. This was demonstrated by labeling mitral and tufted cells with small extracellular horseradish peroxidase injections and processing alternate sections for horseradish peroxidase and for cytochrome oxidase. Because there was cross-reaction of the cytochrome oxidase procedure with horseradish peroxidase, it was possible to trace many neurons through both series of sections. Middle tufted cells of the superficial EPL have basal dendrites confined to the superficial zone of light cytochrome oxidase staining. Internal tufted cells and middle tufted cells of the intermediate zone send their basal dendrites into the intermediate zone. One group of mitral cells (type I) has basal dendrites confined to the deep zone of lighter cytochrome oxidase staining. A second group of mitral cells (type II) and tufted cells of the intermediate EPL has basal dendrites primarily confined to the intermediate zone of dark cytochrome oxidase staining. The correlation of the enzyme staining with the dendritic laminar patterns supports the existence of three relatively distinct sublaminae in the EPL and supports the designation of two types of mitral cells. The staining pattern also provides an independent method for evaluating the sublaminae of the EPL without the necessity of labeling individual groups of cells. Finally, the staining pattern suggests that the intermediate zone of the EPL may be subjected to more tonic synaptic input, causing it to have an increased level of metabolic activity.