Previous studies have shown that damage to vibrissa follicles in newborn rats and mice does not alter the brainstem representations of the remaining vibrissa as demonstrated by staining for mitochondrial enzymes such as cytochrome oxidase (CO) succinic dehydrogenase. This study asked whether this lack of effect might be due to the fact that the trigeminal primary afferents in rodents are already quite well developed at birth. We assessed this possibility by using CO staining the evaluate patterns in the brainstems of pre- and postnatal rats. A vibrissa-related pattern began to emerge in trigeminal nucleus principalis and subnucleus interpolaris (Spl) by embryonic day (E-) 19 and appeared fully developed by the day of birth (P-0). We also made partial lesions of the vibrissa pad on E-15-20 and on P-0, killed pups on P-5-7, and measured the size of the CO-stained patches in Spl on both sides of the brainstem. The correspondence between CO patches and clusters of primary afferent terminal arbors was verified in some animals by combining transganglionic horseradish peroxidase tracing and CO staining. Vibrissa pad damage on E-15-18 resulted in significant (20.1-36.9%) increases in the average area of the remaining CO patches in Spl ipsilateral to the lesion. Vibrissa pad damage on E-19, E-20, and P-0 produced small (6.2-8.9%), but insignificant, increases in patch size in Spl ipsilateral to the lesion. We used anatomical and electrophysiological methods to determine whether our lesions altered the trigeminal innervation of surviving vibrissa follicles. We recorded single trigeminal ganglion cells from 12 rats that sustained vibrissa pad lesion on E-17. As in normal rats, all of the 49 vibrissa-sensitive ganglion cells isolated in the lesioned animals were responsive to deflection of one and only one vibrissa. We also dissected 11 deep vibrissal nerves from intact follicles in adult rats that sustained fetal vibrissa pad damage on E-17, and counted numbers of myelinated axons in 1 microns plastic sections. These data were compared with counts from corresponding follicles on the intact side of the face. The average number of myelinated axons innervating follicles in the damaged vibrissa pads was 196.8 +/- 27.9, and that for the corresponding contralateral nerves was 194.6 +/- 25.7. These data suggest that competitive interactions among the central arbors of trigeminal primary afferents in fetal life may influence the development of central vibrissa representations and, further, that lesion-induced central changes need not be correlated with alterations in the peripheral innervation of undamaged follicles.

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Labelling with 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (Di-A) was used to assess the development of projections within the primary somatosensory cortex (SI) of rats aged between postnatal day 2 and 8 (P-2 and P-8). 1,1'-Dioctadecyl-3,3,3,"3'-tetramethylindocarbocyanine perchlorate (Di-I) was used in these same animals to label thalamocortical afferents. Particular attention was paid to the emergence of lamina IV intracortical projections that form a pattern complementary to vibrissae-related thalamocortical afferents. A vibrissae-related pattern of Di-A-labelled cells and fibers that was restricted largely to the septa regions was not apparent in rats killed on P-2, but it was visible in animals killed on P-4 and later ages. Tracing with biotinylated dextran amine (BDA) was used to assess intra-SI projections of adult rats that sustained transection of the infraorbital nerve (ION) on P-0 or P-7 or implantation of a tetrodotoxin (TTX)-impregnated polymer chip over the cortex on P-0. Rats that sustained ION transection on P-7 or that had TTX implants demonstrated normal patterns of projections within SI. The patterns of labelling in the supra- and infragranular layers of the cortices of the rats that sustained ION transection on P-0 were generally similar to those in the other groups evaluated. However, in lamina IV, there was no organization that could be related to the distribution of the vibrissae. These results indicate that the vibrissae-related pattern of intracortical projections within SI develops shortly after birth and that two manipulations that alter cortical activity, but not the patterning of thalamocortical afferents (application of TTX and transection of the ION after thalamocortical afferent patterns are established), have no significant effect on it. However, a manipulation that alters thalamocortical development (transection of the ION on P-0) profoundly affects the patterning of intracortical connections.

The NaOH sonication digestion technique permits rapid isolation and exposure of intact networks of elastic fibers in vascular tissue for 3-dimensional observation with the SEM. The configuration of the network of elastic fibers within the vascular wall of large elastic arteries (aorta) is generally agreed to be a flexible framework through which smooth muscle cells and collagenous fibers are interwoven. However, the configuration of elastic fiber networks in muscular arteries, medium sized veins and smaller vessels remains unknown. When the lengthy standard biochemical elastin purification techniques were applied to vessels containing lesser amounts of elastic tissue and finer elastic fibers, the vessels were completely digested. In contrast, the digestion and sonication technique isolated and exposed intact networks of delicate elastic fibers in blood vessels which do not contain large amounts of elastic tissue. Unfixed vessels were cut into short segments, placed in 0.5 N NaOH and sonicated for 20-40 min. The specimens were rinsed in deionized distilled H2O, then autoclaved for 30 min. The tissue was rinsed a second time, fixed and processed routinely for SEM. Elastic stains and enzymatic digestion with chromatographically purified elastase and collagenase confirmed that the digestion and sonication technique produced clean, isolated networks of elastic fibers. Knowledge of the configuration of the networks of elastic fibers in different vessels enhances understanding of distensibility characteristics of individual vessels and serves as a baseline for studying alterations in the elastic framework which occur during aging and disease processes such as atherosclerosis, arterial hypertension and aneurysms.

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