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Biopsied skin specimens from 10 males with cutis rhomboidalis nuchae, 50–83 years in age, as well as age-matched non-sun-exposed skin and young normal skin specimens were subjected to a light and electron microscopic immunohistochemical study using anti-elastin polyclonal antibody and anti-microfibril HB-8 monoclonal antibody. Conventional histochemical and electron microscopic techniques were also used. The most severely degenerated elastotic materials had developed in the upper dermis of the cutis rhomboidalis nuchae cases. Light microscopic histochemistry with Elastica van Gieson and Azan staining revealed elastotic materials with elastic fiber characteristics. Electron microscopic observation showed that the elastotic materials consisted of disintegrated and dispersed tannic-acid staining elastin, fine granular materials, and filamentous substances. Light microscopic immunohistochemistry showed that the upper dermis from cutis rhomboidalis nuchae samples was reactive to anti-elastin but not to anti-microfibril HB-8 antibody. Electron microscopic immunohistochemistry further revealed that reticularly disintegrated substances in the elastotic materials retained their reactivity to anti-elastin, but did not show any reactivity to anti-microfibril HB-8 monoclonal antibody. These findings show that these elastotic materials were derived from elastic fibers and partially retained their antigenicity for elastin, but not for microfibrils.

The pacinian corpuscle is composed of an inner and an outer core or bulb. The former is formed by tightly packed and multi-layered thin cellular processes (lamellae) of lamellar cells which surround a centrally located axon terminal, and the latter, also called the capsule, is made up of very loosely piled layers of thin cells which encircle the inner core. Lamellar cells of the inner core are considered to be specialized Schwann cells, and the outer core cells are modified perineurial cells. In the present study, the matrix filling the extracellular spaces of the inner core consisted of basal lamina-like amorphous materials, sparce fine collagen fibrils, and the ground substance embedding these structural components. No definite basal laminae were found on the inner core lamellae except on the peripherally located ones which had distinct basal laminae. Outer-core cells were invested along the entire contour by distinct basal laminae. The interspace between the inner and outer cores was a continuation of the nerve endoneurium. The purpose of this investigation was to determine whether the extracellular matrix of the pacinian corpuscle, especially that of the inner core, has the ability to cause corpuscle regeneration, i.e. to make the regenerating axons and Schwann cells differentiate into corpuscular axon terminals and inner core cells, respectively. Pacinian corpuscles in the periosteum at the distal end of the fibula of mice were repeatedly frozen (3-5 times) in situ with forceps cooled with liquid nitrogen. Within 2-3 days, all the cellular constituents of the corpuscle had degraded, while the extracellular matrices of the inner and outer cores apparently remained undamaged. After 5-7 days, regenerating axons and accompanying immature Schwann cells entered these extracellular matrices of the inner cores. A remarkable finding was that these immature Schwann cells were detached from the axon, and sent thin cellular processes around the axon in a characteristic fashion, basically forming the same pattern as lamellae in a normal corpuscle. The regeneration of the inner core was completed by about 40 days after the freezing treatment. In the outer core, perineurial cells proliferated and extended through the basal lamina tubes of the old cells, becoming new outer core cells. These findings indicate that the extracellular matrix of the pacinian corpuscle has a specific property to cause the regeneration of the corpuscle.

In order to study cell differentiation and morphogenesis of neural retina, ultracytochemical examination for acetylcholine esterase (AChE) was carried out on neural retinal cells from 6-day-old chick embryos cultured in monolayer for 20 days. AChE is a suitable marker for identifying cell specificity and structure of cultured neural retinal cells, because its specific localization in the intact chick neural retina has been established. After about 2 weeks of culturing a number of cell aggregates formed on the monolayer sheet of glial cells, in which cell bodies were generally located on the periphery regions while their cellular processes were in the center, forming neuropil structures. Among such peripherally located cells presumptive ganglion, amacrine, bipolar, and photoreceptor cells could be distinguished. In the neuropil structures, some cellular processes had typical ribbon synapses indicating that these structures correspond to the plexiform layers of the retina. We could also classify the neuropils into two types of both from the AChE activity and from the structure of the nerve terminals. These findings indicate that our cell culture system can be used for the study of cell differentiation and histogenesis of retinal cells.

Nerve segments approximately 6-7 mm long were excised from the predegenerated sciatic nerves of mice, and treated 5 times by repetitive freezing and thawing to kill the Schwann cells. Such treated nerve segments were grafted into the original place, being in contact with the proximal stump of the sciatic nerve. The animals were sacrificed 2, 3, 5, 7 and 10 days, 2, 3, 5 and 8 weeks after the grafting. The grafts were examined at the middle level, i.e., about 3-4 mm distal to the proximal end of the graft, by light and electron microscopy. Within 2-3 days after the grafting, the dead Schwann cells were disintegrated into fragments and gradually phagocytized by macrophages. However, the basal laminae of the Schwann cells remained as empty tubes (basal lamina scaffolds). The notable finding was that the regenerating axons always grew through these basal lamina scaffolds. New Schwann cells seemed to migrate along these axons from the proximal stumps. The number of axons growing through the basal lamina scaffolds gradually increased with time. These axons were surrounded in a bundle by Schwann cells. About 1 week after the grafting, axons began to be segregated into smaller bundles by Schwann cells. Axons with a relatively large diameter (about 2 microns) tended to be sorted out and surrounded by their own Schwann cells. The myelination began about 2 weeks after the grafting on such large diameter axons. The basal lamina scaffolds, through which the regenerating axons had grown, were gradually disintegrated into fragments by the expansive forces due to the increase in number and volume of the regenerating axons and Schwann cells. Groups of axons, which had been derived from the same basal lamina scaffolds, were enclosed with the cells resembling perineurial epithelial cells. These perineurial epithelial cells proliferated and further separated groups of axons into smaller ones or even into single axons. The number of myelinated axons increased with the advancement of regeneration. These results show that the basal lamina scaffolds of Schwann cells serve as efficient conduits for the elongation, maintenance and maturation of regenerating axons.

✓ In order to examine nerve regeneration under conditions in which the basal laminae of the glial limiting membranes (GLM) and blood vessels were preserved intact, the intraorbital segment of adult rat optic nerve was frozen locally. During the next 3 months, degenerative and regenerative changes in axons and glial cells were observed by light and electron microscopy. On the day after treatment, all the myelinated and unmyelinated axons in the central zone of the lesion were damaged. The astrocyte endfeet of the GLM as well as the blood vessels were extensively disrupted, while their basal laminae were preserved apparently intact as a continuous sheet. Three days after treatment, regenerating axons appeared in the central zone of the lesion. They contained various numbers of clear and dense-cored vesicles as well as some smooth endoplasmic reticulum. The regenerating axons gradually increased in number, especially beneath the pial and perivascular surfaces of the lesion, where an abundance of regenerating axons was found 3 months after treatment. A few of these axons were abnormally remyelinated by oligodendrocytes. In addition to this axonal regeneration through the intraoptic nerve compartment, fine regenerating axons were seen growing out through GLM into the pial connective tissue 3 weeks after treatment. Astrocyte endfeet of the GLM became irregular in contour, protruding in a fern-leaf fashion into the pial connective tissue. Fine naked axons grew out through these protrusions and subsequently increased in number, vigorously growing in large bundles both proximally and distally along blood vessels in the pial connective tissue. Bundles of regenerating axons extended as much as 1.5 mm from the site of the lesion 3 months after surgery. These bundles were covered by thin processes of pial or arachnoidal non-neuronal cells, and the regenerating axons remained unmyelinated. The above findings indicate that under well-nourished conditions, adult mammalian optic nerve exhibits considerable regenerative ability.

Nerve endings of the trigeminal nerve were examined in the lip, nostril, upper jaw and supraorbital skin of cat by the horseradish peroxidase (HRP) tracing method. Wheatgerm agglutinin-HRP (WAGA-HRP) as injected into the spinal trigeminal nucleus was transported transganglionically to nerve endings. The present study deals with presumptive baroreceptive nerve endings found on arteriovous anastomoses and on some other large arteries. By light microscopy HRP-labeled, complexly arborized nerve endings were found on the walls of arteries located deep in the subcutaneous tissue of the lip and nostril. These arteries were identified in serial sections to be an arterial segment of an arteriovenous anastomosis. By electron microscopy characteristic nerve endings were located in the adventitia and partly extended into the adjacent connective tissue of the arterial wall. These terminals had no connective tissue capsule. Axon terminals were somewhat enlarged (1–2μm in diameter) and extended through bundles of collagen fibrils. The terminals contained an abundance of mitochondria and some vesicles. Axon terminals were typically covered by thin Schwann cell processes, but parts of the axolemma were sometimes devoid of such Schwann cell coverings, being invested only by basal laminae. Cell bodies of Schwann cells were located apart from axons. These light and electron microscopic features of the endings resembled those of other well-defined baroreceptors reported in the carotid sinus, aortic arch and endocardium, as well as of Ruffini terminals and Golgi tendon organs, suggesting that they would be baroreceptors of arteriovenous anastomosis. In addition HRP-labeled single nerve fibers with varicosities were found in the walls of some large arteries in the facial skin. By electron microscopy, such a HRP-positive nerve fiber contained some mitochondria and vesicles in varicosities and coursed with a bundle of HRP-negative fine fibers in the adventitia of arteries. These HRP-labeled single fibers were considered to be sensory derived from trigeminal nerves.

The Pacinian corpuscle consists of an axon terminal and specialized non-neuronal cells of an inner and an outer core (Pease and Quilliam, 1957; Nishi et al., 1969; Munger, 1971; Halata, 1977; Chouchkov, 1978; Malinovsky and Pac, 1982). The inner core is formed by densely packed thin cytoplasmic processes (lamellae) of lamellar cells which surround the axon terminal in a bilaterally symmetrical fashion. These inner core lamellar cells are thought to be derived from Schwann cells. There are thin spaces between neighboring lamellae, in which an extracellular matrix including basal lamina-like material and fine collagen fibrils is present. The outer core is composed of loosely piled layers of thin cells concentrically encircling the inner core. There are definite basal laminae on these outer core cells and thin processes. The outer core cells are extensions of the perineural epithelial cells of the innervating nerve (Shanta and Bourne, 1968). The axon terminal is characterized by an accumulation of mitochondria beneath the axolemma and by small cytoplasmic processes (axonal spines) emanating from the axolemma (Munger et al., 1987; Ide et al., 1987a). It has been demonstrated that basal laminae can serve as conduits for regenerating axons in the peripheral nerve (Ide et al., 1983; Osawa et al., 1986). As described above there are extracelluar matrices containing basal lamina-like material in the inner core of Pacinian corpuscles.

Summary Part of the dorsal funiculus of the adult male rat (Wistar) spinal cord was treated for l h at the thoracolumbar level by running hot water, at approximately 48–50 ° C, through a polyethylene tube 2 mm in diameter in contact with the dura. Animals were fixed 1 day to 4 weeks later and the spinal cords were examined by light and electron microscopy. The affected area in the dorsal funiculus was approximately 1 mm long and less than 1 mm wide at the dorsal surface, and varied from 0.4 to 0.7 mm in depth. Within 3 days after treatment, almost all the myelin sheaths in the affected area were degraded, leaving the axons denuded, and at the same time astrocyte endfeet at the glial limiting membrane were swollen and partly destroyed. Almost all the denuded axons remained intact, exhibiting no noticeable morphological changes. There was evidence of a moderate vasogenic oedema, but minimal signs of haemorrhage in the lesion. Seven days after treatment, many immature Schwann cells but no oligodendrocytes were found between the denuded axons. By 2 weeks many of the denuded axons were remyelinated, and by 4 weeks almost all of those axons located near the pial and perivascular surfaces had been remyelinated by Schwann cells, while most of those located in the deep and marginal zones bordering the adjoining intact areas were remyelinated by oligodendrocytes. Longitudinal sections revealed that at nodes of Ranvier PNS-type myelin sheaths were apposed by either intact or newly formed CNS-type myelin sheaths. A typical glial limiting membrane was not reformed beneath the pial surface, but an inconspicuous one was found between the PNS- and CNS-type fibre areas.

The distribution of carbonic anhydrase (CA) activity was studied by electron microscopic histochemistry in rat Pacinian corpuscles, Meissner corpuscles and Merkel cell-neurite complexes using the cobalt bicarbonate method. The distribution of CA activity in these axon terminals was compared to the activity in sciatic nerve axons. An intense enzymatic CA activity was demonstrated in axon terminals of both Pacinian and Meissner corpuscles, while a weak activity was found within the axoplasm of terminals abutting Merkel cells. Some large- and medium-sized axons in sciatic nerves exhibited an intense activity. These findings indicate that large- or medium-diameter sensory axons innervating corpuscular endings have an intense CA activity extending from their somata to their sensory terminals. Axons to Merkel-neurite complexes differ in CA activity from those innervating Meissner and Pacinian corpuscular endings.

The ultrastructure of cat mesenteric Pacinian corpuscles in cross and longitudinal sections has been examined. The terminal ends of lamellar cells of the inner core have been identified in longitudinal sections through the proximal portion of the inner core. These terminal bulbous expansions contain characteristics concentric membranes of rough endoplasmic reticulum and in some cases masses of oval membranous inclusions. The central axon as seen in cross section is oval in profile, having X-(short) and Y-(long) axes, and each axonal face is characterized by specializations of the axolemma. At the X-axis, the inner lamellae of the inner core tightly abut a smooth axolemma, with no intervening connective tissue matrix, in a manner reminiscent of a neuroepithelium. The axolemma of the Y-axis has numerous axonal spines (microspikes) that project into the cleft in the inner core. The extent of the axolemma having axonal spines can only be appreciated in longitudinal sections. The clefts contain a specialized connective tissue with elastic and collagen fibrils. The connective tissue compartment of fibers and matrix separating individual inner core lamellae is unique, in that it contains extremely thin collagen fibrils measuring approximately 15 nm in diameter. The diameter of collagen fibrils increases as the cleft is approached. Here the fibrils resemble typical endoneural collagen.

The present review of cutaneous sensory receptors begins with a consideration of free nerve endings (FNEs) that can be considered as sensory terminals evidencing the least structural specialization of the axon and associated cells. Using the criteria established by KRUGER et al (1981), FNEs of both A delta and C fibers can be identified on the basis of ultrastructural characteristics that include an intimate relationship between axons and the associated epithelium, the lack of a complete Schwann cell investment, the accumulation of numerous vesicles and other cytoplasmic organelles, and for A delta terminals a 1:1 relationship between axon and investing Schwann cell. Using these criteria, the so-called genital end bulbs of the human glans penis are merely a skein of FNEs based on the ultrastructural study of HALATA and MUNGER (1986).Hair follicles of most species studied to date (the exception being the rabbit and to some extent the guinea pig) are multiply innervated with lanceolate, Ruffini and FNEs. The lanceolate terminals are the rapidly adapting terminals that are numerous in guard hairs. Ruffini terminals of hairs resemble those of the periodontal ligament or joint capsules and both are remarkably similar to Golgi tendon organs in terms of ultrastructural characteristics. The key ultrastructural characteristic is the encircling of collagen bundles by axons and associated Schwann and connective tissue cells.Axons frequently enter the epidermis either to terminate as FNEs or become associated with Merkel cells in glabrous skin at the base of the papillary ridges or in clusters of Merkel cells in hairy skin in touch domes or Haarscheiben. Merkel cells have clusters of apparent secretory granules polarized toward the axon and the axon is typically a slowly adapting mechanoreceptor. The function of the granules is not known.Pacinian corpuscles are the largest of the corpuscular receptors of the dermis and are characterized by an elaborate inner core of stacks of numerous thin lamellae arranged in a bilaterally symmetrical manner. Based on the fact that the lamellae are coupled with gap junctions and the outer core lamellae isolated by numerous tight junctions, the authors have proposed that the unique ionic environment may be in part responsible for the remarkable sensitivity of Pacinian corpuscles (MUNGER and IDE, 1987).Meissner corpuscles are a typical corpuscular receptor of murine (IDE, 1976, 1977), marsupial and primate glabrous skin (MUNGER, 1971). The axons typically weave back and forth between stacks of lamellae. Simple corpuscles are common in many mammals especially in the glabrous snout skin (MUNGER, 1965, HALATA and MUNGER, 1983). The lamellae around the axon are typically arranged circumferentially around the axon.The authors have previously noted that axons in mechanoreceptors including lanceolate terminals, Pacinian and Meissner corpuscles, and muscle spindles frequently have axonal spines projecting from the elongated or Y-axis of the axon (MUNGER and IDE, 1987). Axonal spines typically are filled with filaments and lack other cytoplasmic organelles. The common presence of axonal spines provided the basis for the speculation that they may be involved in mechano-electric transduction.

The present report is the second of two studies re-evaluating the cytological characteristics of Pacinian corpuscles. The extreme tip of the axon of a Pacinian corpuscle has been identified and is quite different from the previously described ultraterminal region. The latter is the site where the inner core lamellae begin to terminate and is characterized by a smooth axolemma. The extreme tip lacks inner core lamellae directly abutting the axolemma and is instead characterized by the presence of many axonal spines projecting into a matrix of basal lamina-like material. The extreme tip of the axon thus resembles the organization of the axolemma facing the clefts of the inner core. The axonal spines at the cleft and extreme tip are proposed as a site of restricted current flow due to the tight apposition of inner core lamellae to the axolemma of X-axis. The hemi-inner cores thus could restrict current flow to the cleft. These anatomical specializations could represent both a source and a sink for K+ ions during mechano-electric transduction and account in part for the exquisite sensitivity of Pacinian corpuscles to complex pressure waves.