Localization and distribution of laminin in the basal lamina of certain mechanoreceptors—an immunogold study

The applied immunogold cytochemical technique in investigating the cytologic distribution of the laminin (LAM) molecule in the capsulated Pacinian and Herbst mechanoreceptors shows the presence of LAM around most elements of the receptor structures. The LAM immunoreactivity (LAM-IR) is best expressed in the vicinity of the perineural capsule cells of both receptor types, where it is primarily concentrated around the perinuclear regions as well as the cytoplasmic lamellae. Such a localization overlaps with the already known ultrastructural localization of a basal lamina (BL) around these cells. Laminin immunoreactivity is less well expressed around the modified Schwann cells. Even in these cells, however, there is an apparent immunoreaction around the cytoplasmic lamellae regardless of the lamellar location. In both receptor types, there is no LAM-IR in the cells of the subcapsular space. Of particular significance we consider the localization of gold particles (respectively the presence of a BL) between the innermost lamellae of the modified Schwann cells and the non-myelinated part of the receptor nerve fiber and their endings, as well as around the axoplasmic protrusions of the nerve endings. We discuss the role of the BL and LAM in the investigated rapidly adapting mechanoreceptors and their trophic influence upon the sensory regions. We also assume the arresting and selective effect of these membranes in building up the ion channels of the axolemma which probably has a certain importance in mechanotransduction.     

Fine-Structural Characterization of the Somatic Innervation of the Tympanic Membrane in Normal,Sympathectomized, and Neurotoxin-Denervated Rats

The pars tensa of the rat tympanic membrane (TM) consists largely of a lamina propria of specialized unbanded collagen bounded on the outer surface by an unusually thin epidermal layer and on the inner surface by a flat, single-cell mucosal layer. The mucosal layer is innervated solely by unmyelinated (C) axons, whereas the cutaneous layer is supplied by both myelinated and C axons. The outer surface differs from general body skin, lacking dermal papillae, hairs, sweat glands, and distinctive dermal corpuscular structures.

Epidermal innervation includes distinctive terminals in the basal layer, unassociated with Merkel cells, and deeper intraepidermal smaller endings containing accumulations of mitochondria and vesicles. The sensory nature of these endings can be inferred by their extensive, but not total, elimination following neonatal capsaicin treatment (a potent neurotoxin for thin sensory fibers) and their preservation following surgical or neurotoxin sympathectomy.

The thin mucosal epithelium displays capillaries and beaded axons close to the free surface of the middle ear. The unmyelinated terminals contain predominantly large, dense-core vesicles (LDCVs). Capsaicin treatment results in extensive elimination of terminals containing LDCVs in surface epithelia. A possible small trophic influence of sensory thin-fiber supply was noted on the development of the epidermal layers. The sensory modalities elicited by natural stimulation of the TM is considered in relation to the pattern of innervation.     

Immunocytochemical identification of proteins within the Pacinian corpuscle

Light- and electron-microscopic immunocytochemistry (ICC) was performed on Pacinian corpuscles (PCs) obtained from cat mesentery to determine the presence and location of various proteins within the accessory capsule and the neurite. Antibodies to tubulin, neurofilament 200, actin, collagen II and V, glial fibrillary acidic protein (GFAP) and S-100 were used. Type II collagen was localized only in the outer core of the accessory capsule, which is composed of an inner core, an intermediate layer or growth zone, an outer core and an external capsule. Type V collagen was found only in the intermediate growth zone. Intermediate filaments labeled with anti-GFAP were only found in the inner core. The calcium-binding protein that was labeled by anti-S-100 was found only in the inner core. Diffuse and variable staining for actin is present throughout the accessory capsule. The differences in distribution of these various proteins within the capsule suggest different structural/functional properties of the various capsule regions. The neurite was found to contain microtubules (i.e., tubulin) and neurofilaments throughout, but these cellular inclusions were not found within the cytoplasmic extensions (filopodia) that project from the neurite into the hemilamellar clefts formed by the inner-core hemilamellae. The extensions, however, were found to contain actin in a much greater density than that seen in the neurite proper. The presence of actin, but apparent lack of other cytostructural elements within the extensions, is highly reminiscent of the composition of stereocilia found on vestibular and auditory hair cells. Since stereocilia have been shown to play a role in hair-cell mechanotransduction, it is possible that the cytoplasmic extensions are significantly involved with mechanotransduction within the PC.     

Immunohistochemical study of cutaneous nerves in the emu

The distribution and chemical content of cutaneous nerves in 3- to 13-day-old emu chicks (Dromaius novaehollandiae) were examined by using double-labelling immunohistochemistry. Seven different subpopulations of cutaneous nerves were identified based on their neurochemistry. No intraepidermal nerve fibres were found. However, axons were located within the dermis and were often associated with blood vessels, pennamotor muscles and feather follicles or innervated Herbst corpuscles. Both similarities and differences exist between subpopulations of cutaneous nerves in the emu and volant birds. As in volant birds, a subpopulation of cutaneous axons innervates the superficial skin layers and contains immunoreactivity to both substance P and calcitonin gene-related peptide (CGRP). This suggests that the neuropeptide content of these presumptive free nerve endings is conserved throughout the evolution of birds. In contrast, Herbst corpuscles in the emu are innervated by axons that contain immunoreactivity for CGRP or neuropeptide Y (NPY) but that lack the calbindin D-28k immunoreactivity found in fibres innervating Herbst corpuscles of volant birds. Herbst corpuscles therefore may have a different chemical content in a flightless species from that in volant birds.This work was supported by an Australian Postgraduate Award (Industry; CO9942100) and the Flinders Institute for Health and Medical Research.     

Tactile sensitivity of normal and autistic children

Many children with autistic spectrum disorders have unusual reactions to certain sensory stimuli. These reactions vary along a hyper- to hypo-responsivity continuum. For example, some children overreact to weak sensory input, but others do not respond negatively to even strong stimuli. It is typically assumed that this deviant responsivity is linked to sensitivity, although the particular stage of sensory processing affected is not known. Psychophysical vibrotactile thresholds of six male children (age: 8–12) who were diagnosed to have autistic spectrum disorders and six normal male children (age: 7–11) were measured by using a two-alternative forced-choice task. The tactile stimuli were sinusoidal displacements and they were applied on the terminal phalanx of the left middle finger of each subject. By using a forward-masking paradigm, 40- and 250-Hz thresholds of the Pacinian tactile channel and 40-Hz threshold of the Non-Pacinian I tactile channel were determined. There was no significant difference between the thresholds of autistic and normal children, and the autistic children had the same detection and masking mechanisms as the normal children. The sensory responsivity of each subject was tested by clinical questionnaires, which showed again no difference between the two subject groups. Furthermore, no significant correlations could be found between the questionnaire data and the psychophysical thresholds. However, there was a high correlation between the data from the tactile and emotional subsets of the questionnaires. These results support the hypothesis that the hyper- and hypo-responsivity to touch, which is sometimes observed in autistic spectrum disorders, is not a perceptual sensory problem, but may probably be emotional in origin.     

The Effect of Stimulus Duration on Frequency-Response Functions in the Pacinian (P) Channel

Psychophysical experiments on human observers and physiological measurements on Pacinian corpuscles (PCs) isolated from cat mesentery were performed to explain certain discrepancies in the psychophysical—physiological model (Bolanowski et al., 1988) for the sense of touch in the vibrotactle Pacinian (P) channel. The model was based on correlations among the psychophysical frequency response obtained on human glabrous skin and physiological frequency-response functions measured on two PC preparations: PC fibers innervating human glabrous skin (Johansson et al., 1982) and PCs isolated from cat mesentery. The three frequency-response functions were qualitatively similar. However, the low-frequency slope for the human PC fibers differed from the slopes for the psychophysical and cat mesentery PC functions by being 3 dB/octave less steep. This discrepancy can be explained theoretically by differences in methodology involving the effect of stimulus duration and the property of temporal summation known to exist in the P channel (i.e., a 3-dB increase in sensitivity per doubling of stimulus duration). To test this, experiments were performed using two methods of stimulation: (1) a constant stimulus duration for different test frequencies, as generally used in this laboratory; and (2) a constant number of stimulus cycles (n = 5) for each test frequency as used by Johansson et al. The method of least squares was used to calculate the low-frequency (50 to 150-Hz) slopes of individual psychophysical and physiological functions. The mean slopes that resulted from using the two methods of stimulation were consistent with the theoretical expectations.     

The Postnatal Functional Development of Muscle Stretch Receptors in the Rat

The response to a 5-sec stretch of the triceps muscle was studied in dorsal root filaments L₅ of 72 infant rats (1-19 days old) under urethane anesthesia.

More than 50% of all units in 1-day-old rats responded by repetitive firing until the end of the 5-sec stretch (slowly adapting or SA receptors), while the rest ceased to fire earlier (relatively rapidly adapting or l/2 SA receptors), or gave an “on” response only. The number of units exhibiting an SA response increased with age and attained 80% in 5-day-old rats. By the 10th day of life, almost 90% of endings behaved as SA receptors. During development, the maximal discharge frequencies at the peak of stretch increased markedly, and their values in 18-day-old rats were comparable to those in adult rats. The phasic component of the response to stretch, although less well defined in the younger animals, was already present even in 1 -day-old rats. Adaptation of the static response during maintained stretch was relatively steep in all the age groups studied.

The results indicate that, in the rat, large numbers of muscle stretch receptors are capable of responding to sustained stretch as SA receptors, even at an age when their morphological and ultra-structural maturation is not yet fully accomplished.     

Immunoblot findings of calcareous corpuscles binding proteins in cyst fluid of Taenia solium metacestodes

After collecting calcareous corpuscles from plerocercoid of Spirometra mansoni (sparganum), we evaluated the antigenic values of calcareous corpuscles binding proteins obtained from the cyst fluid of Taenia solium metacestodes. Immunoblot analysis revealed that cysticercosis patient sera strongly recognized 10 and 95 kDa calcareous corpuscles binding proteins. This result demonstrated that calcareous corpuscles are bound with major secretory antigenic proteins, which is possibly involved in the secretory pathways of the 10 and 95 kDa proteins presenting in the cyst fluid of T. solium metacestodes.     

Cytokeratin expression in human thymus: immunohistochemical mapping

Cytokeratin expression in normal postnatal human thymus was studied immunohistochemically by using monoclonal antibodies against various cytokeratin polypeptides. An attempt was made to characterize cell populations giving rise to the cornified structures of Hassal's corpuscles. Monoclonal antibody KB-37, a marker of squamous epithelium basal cells, was applied to distinguish the earliest cells capable of undergoing squamous differentiation. Parts of the subcapsular epithelium were extensively stained with this reagent. This epithelium, like the basal layer of certain squamous epithelia, exibited a high incidence of cytokeratins 13 and 14, and pronounced expression of cytokeratin 19. Simple epithelium cytokeratins 8, 18, and 19 were present in the cortex. Scattered cells reacted with KB-37 antibody. All stellate epithelial cells in the medulla were positive for cytokeratin 19. Most of the medullar epithelial cells were positive for cytokeratins 13, 14 and 17 of complex epithelium, in contrast to the cortex, where only a few cells were positive for these cytokeratins. A significant proportion of the medullar cells was positive for KB-37 antigen. Cytokeratins 8 and 18 were expressed in single cells and in groups of cells surrounding Hassal's corpuscles. The outermost cells of these corpuscles were positive for cytokeratin 19 and KB-37. In the peripheral parts of Hassal's corpuscles, simple epithelium cytokeratins 7, 8, 18, and cytokeratins 4, 13, 14, and 17, characteristic of stratified nonkeratinizing epithelia, were coexpressed with keratinization-specific cytokeratins 10/11. The inner parts of the swirls were uniformly positive for cytokeratins was reduced.     

Structure of the kidney in the coelacanth Latimeria chalumnae with reference to osmoregulation

The morphology of the nephrons of the coelacanth Latimeria chalumnae was investigated by light microscopy. Each nephron is composed of a large renal corpuscle with well‐vascularized glomerulus, non‐ciliated neck segment, proximal convoluted tubule divided into distinct first and second segments, non‐ciliated intermediate segment, distal tubule, collecting tubule and collecting duct. The parietal layer of the Bowman's capsule of the renal corpuscle is composed of low cuboidal cells. The short non‐ciliated neck segment is lined by cuboidal epithelium. The first and second proximal segments display a prominent brush border and contain amorphous material in their lumen. The second proximal segment differs from the first segment in having taller columnar epithelium and a relatively narrow lumen. The intermediate segment is lined by non‐ciliated columnar epithelium and its lumen appears empty. The distal tubule is narrow in diameter and its cuboidal epithelium is devoid of intercalated cells. A unique feature of L. chalumnae is having binucleate cells in the tubule and collecting duct epithelium. The renal arteries have poorly developed tunica media and its cells contain granular material. The structure of L. chalumnae nephrons correlates well with their osmoregulatory function and resembles those of euryhaline teleosts.