Improved contrast in histochemical detection of cytochrome oxidase: metallic ions protocol.

The standard current technique for demonstration of cytochrome oxidase (CyOx) provides low-contrast diaminobenzidine (DAB) polymer. In order to enhance the contrast with divalent metalic ions, we have screened a number of buffers and found that Hepes, Mops and cacodylate neither precipitate these ions nor inactivate CyOx in a concentration of 0.1 M. Staining thus obtained shows a broad range of gradations between black and white. With fresh tissue the resulting image is superior to that obtained with the brown DAB product, even if a recommended blue filter or printing on very hard paper are used. The technique is as simple as the one which is currently standard. Fixed tissue, cut either in a cryostat or vibratome, can be stained well when floating but not when mounted on slides. The stained floating tissue can be used for electron microscopy, but has no advantage over the standard method.

Focal cortical seizures prevent HRP and HRP-WGA labeling only in neurons bidirectionally connected to the cortex.

Intracortical implants of polyacrylamide gel containing horseradish peroxidase labeled cortical efferents and perikarya in some cortical areas and a number of subcortical formations. When epileptogenic penicillin was added to the gel, no labeling was seen in the efferents and cell bodies of the cortex, thalamus, or claustrum, whereas the magnocellular nuclei of the basal forebrain, raphe nuclei and locus coeruleus did contain the label.

Different metabolic changes in the lateral geniculate nucleus and the superior colliculus of adult rats after simultaneous or delayed double enucleation.

By means of the [14C]deoxyglucose method the local cerebral glucose utilization (LCGU) was measured in the lateral geniculate nucleus and the superior colliculus of rats with a simultaneous double enucleation, animals with an enucleation of the right eye between neonatal and adult stages followed by an enucleation of the left eye some months later and adult control rats. The control animals show LCGU values within the limits published by other observers. The LCGU values of the simultaneously double-enucleated rats are reduced on both sides to the same extent by about 20%. The dorsal lateral geniculate nucleus shows the largest decrease. The LCGU values of the rats with the delayed double enucleation have decreased even more (up to 37%). The right sided visual brain regions have a significantly lower LCGU than the corresponding regions on the left side. These findings indicate that unilateral enucleation from neonatal to adult stages leads to adaptive changes resulting in a higher metabolic vulnerability, which is revealed by the second enucleation.

Cortical projections of the thalamic mediodorsal nucleus in the rat. Definition of the prefrontal cortex.

Deposits of somatopetal tracers, that filled the entire dorsolateral cortex of one hemisphere, labelled only a few cell bodies in the thalamic mediodorsal nucleus (MD). Further experiments served to confirm the existence of strong projections from MD to the mesial and suprarhinal areas of the frontal cortex. Thus, only a very small proportion of the MD neurones in the rat projects outside the cortical areas described first by Leonard (1969). We conclude that at least in the rat, cortical projections of the MD are selective enough to be used as the basis for definition of the prefrontal cortex.

Absence of transsynaptic transport in cerebello-thalamo-cortical path of the rat.

Attempts to visualize the cerebello-thalamo-cortical path in the rat were made with different approaches. We tried (1) double labelling with somatopetal tracing from the motor cortex and somatofugal from the cerebellar nuclei, (2) transmembrane labelling by depositing biocytin or wheat germ agglutinin (WGA) into the motor cortex or cerebellum. WGA was either iodinated with 125I or conjugated with horseradish peroxidase (HRP). The double labelling technique showed an overlap of the tracers in the same thalamic region but no evidence of transsynaptic transport in either direction was obtained. Our results indicate a difference in the organization of this system in primates and rodents, since transsynaptic labelling in the cerebellar nuclei after injections of WGA-HRP conjugate in the monkey motor cortex has been found.