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1.
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.  相似文献   

2.
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.  相似文献   

3.
This investigation presents new insights into the structure of human secondary lamellae. Lamellar specimens that appear dark and bright on alternate osteon transverse sections under circularly polarizing light were isolated using a new technique, and examined by polarizing light microscopy, synchrotron X-ray diffraction, and confocal microscopy. A distribution of unidirectional collagen bundles and of two overlapping oblique bundles appears on circularly polarizing light microscopy images in relation to the angle between the specimen and the crossed Nicols' planes. The unidirectional collagen bundles observed at 45 degrees run parallel to the osteon axis in the dark lamellar specimens and perpendicular to it in the bright ones. Small and wide-angle micro-focus X-ray diffraction indicates that the dark lamellae are structurally quite homogeneous, with collagen fibers and apatite crystals preferentially oriented parallel to the osteon axis. Bright lamellar specimens exhibit different orientation patterns with the dominant ones bidirectional at +/-45 degrees with respect to the osteon axis. Accordingly, confocal microscopy evidences the presence of longitudinal bundles in dark lamellar specimens and oblique bundles in the bright ones. Radial bundles are evidenced in both lamellar types. The alternate osteon structure is described by a rather continuous multidirectional pattern, in which dark and bright lamellae display different mechanical and possibly biological functions.  相似文献   

4.
5.
Spiral ganglion neurons (SGNs) provide afferent innervation to the cochlea and rely on contact with hair cells (HCs) for their survival. Following deafferentation due to hair cell loss, SGNs gradually die. In a rat culture model, we explored the ability of prosurvival members of the Bcl-2 family of proteins to support the survival and neurite outgrowth of SGNs. We found that overexpression of either Bcl-2 or Bcl-xL significantly increases SGN survival in the absence of neurotrophic factors, establishing that the Bcl-2 pathway is sufficient for SGN cell survival and that SGN deprived of trophic support die by an apoptotic mechanism. However, in contrast to observations in central neurons and PC12 cells where Bcl-2 appears to promote neurite growth, both Bcl-2 and Bcl-xL overexpression dramatically inhibit neurite outgrowth in SGNs. This inhibition of neurite growth by Bcl-2 occurs in nearly all SGNs even in the presence of multiple neurotrophic factors implying that Bcl-2 directly inhibits neurite growth rather than simply rescuing a subpopulation of neurons incapable of extending neurites without additional stimuli. Thus, although overexpression of prosurvival members of the Bcl-2 family prevents SGN loss following trophic factor deprivation, the inhibition of neurite growth by these molecules may limit their efficacy for support of auditory nerve maintenance or regeneration following hair cell loss.  相似文献   

6.
We studied effect of gangliosides on viability of brain neurons and neuronal PC12 cell line exposed to toxic concentrations of compounds activating free radical reactions. It is found that preincubation of cerebellar granule cells and PC12 cells with micromolar concentrations of ganglioside GM1 increases statistically significantly viability of these cells submitted to inductors of oxidative stress, such as hydrogen peroxide and the Fe2+-ascorbate system However, the effect of ganglioside GM1 in the PC12 cells failed to be revealed 1–2 days after treatment of the cells with trypsin, which indicates an importance of interaction of gangliosides with surface proteins for realization of their protective action. GM1, GD1a, and other gangliosides were shown to produce the neuroprotective effect on cerebellar granule cells in the presence of toxic glutamate concentrations. Not only micro-, but also nanomolar concentrations of these gangliosides increased statistically significantly the neuronal viability, although at micromolar concentrations this effect as a rule was more pronounced. The obtained data allow suggesting that the neuroprotective action of gangliosides is determined to a considerable degree by their ability to inhibit free-radical reactions in nerve cells.  相似文献   

7.
Book review     
It has long been accepted that action potentials arising from Pacinian corpuscles (PCs) originate at the first node of Ranvier located within the PC and that the mechanotransduction events (receptor potentials) are formed by stretch-activated channels selectively sensitive predominantly to Na +. Also, it has been shown previously that tetrodotoxin (TTX) affects the receptor potential suggesting that transduction may involve voltage-sensitive Na + channels. To determine whether voltage-sensitive Na + channels exist in the membrane thought to be responsible for transduction, immunocytochemical studies were performed using polyclonal antibodies raised in rabbit against the alpha subunit of rat type I and type II voltage-gated sodium channels. The results show the presence of label on the neurite and axolemma, as well as in the node regions. Interestingly, labeling is also found on the inner and outer lamellae that form the non-neural accessory structure surrounding the neurite. The presence of this label in the surrounding lamellae suggests that voltage-sensitive Na + channels, that are involved in both transduction and action-potential generation, may be made available to the neurite via transport from the lamellae, a mechanism perhaps operating in parallel to axoplasmic transport.  相似文献   

8.
Previously we found that some cyclopentenone prostaglandin derivatives promoted neurite outgrowth from PC12 cells and dorsal root ganglia explants in the presence of nerve growth factor; and so we referred to them as neurite outgrowth-promoting prostaglandins (NEPPs). In this study, NEPPs protected HT22 cells against oxidative glutamate toxicity. NEPP6, one of the most effective promoters of neurite outgrowth in PC12 cells, protected the cells most potently among NEPPs 1--10. Several derivatives, NEPPs 11--19, were newly synthesized based on the chemical structure of NEPP6. NEPP11 had a more potent neuroprotective effect than NEPP6. NEPP11 also prevented the death of cortical neurons induced by various stimuli and reduced ischemic brain damage in mice. Biotinylated compounds of NEPPs were synthesized to investigate their cellular accumulation. NEPP6-biotin protected the cells and emitted potent signals from the cells. In contrast, biotinylated non-neuroprotective derivatives emitted much weaker signals. These results suggest that NEPPs are novel types of neurotrophic compounds characterized by their dual biological activities of promoting neurite outgrowth and preventing neuronal death and that their accumulation in the cells is closely associated with their neuroprotective actions.  相似文献   

9.
It has long been accepted that action potentials arising from Pacinian corpuscles (PCs) originate at the first node of Ranvier located within the PC and that the mechanotransduction events (receptor potentials) are formed by stretch-activated channels selectively sensitive predominantly to Na+. Also, it has been shown previously that tetrodotoxin (TTX) affects the receptor potential suggesting that transduction may involve voltage-sensitive Na+ channels. To determine whether voltage-sensitive Na+ channels exist in the membrane thought to be responsible for transduction, immunocytochemical studies were performed using polyclonal antibodies raised in rabbit against the alpha subunit of rat type I and type II voltage-gated sodium channels. The results show the presence of label on the neurite and axolemma, as well as in the node regions. Interestingly, labeling is also found on the inner and outer lamellae that form the non-neural accessory structure surrounding the neurite. The presence of this label in the surrounding lamellae suggests that voltage-sensitive Na+ channels, that are involved in both transduction and action-potential generation, may be made available to the neurite via transport from the lamellae, a mechanism perhaps operating in parallel to axoplasmic transport.  相似文献   

10.
Summary The ultrastructure of Pacinian corpuscles of the cat located in the crural region and innervated by the interosseous nerve was studied 1 to 14 months after denervation. Both the Pacinian inner core and capsule remained well preserved one month after denervation. However, the denervated inner cores underwent progressive atrophy and wasting, which resulted in a gradual reduction of the amount of inner-core cells and lamellae, widening of interlamellar clefts, formation of empty spaces in the axial region and a considerable increase in the number of collagen fibrils. In spite of the wasting, the inner core still survived 14 months after denervation, but at least half of its volume became occupied by collagen fibrils which surrounded the remaining inner-core cells and lamellae. Collagen fibrils assembled in the denervated core were markedly thinner than those found in the capsule, as is also the case in normal Pacinian corpuscles. In the capsule, discrete focal degeneration, occasional pyknosis of the innermost capsular cells and macrophage infiltration were observed from the first month after nerve section onward, but the number of capsular layers remained within the normal range (30–40) up to 14 months after denervation.  相似文献   

11.
The aim of the present study was to investigate the responses of Merkel cells that are numerous in the palatine rugae, due to the continuous mechanical stimulation exerted by the palatal plate. Forty golden hamsters were used in this experiment. The palatal plate was made of adhesive resin and it was set on the palate of the animal. To exert a continuous pressure, a 0.8 mm elevation on the internal surface of the palatal plate was created at the middle portion of the fourth palatine ruga. Thereafter, the number of Merkel cells in the mucosa was calculated by immunohistochemical observation. Morphological changes of Merkel cells were examined by electron microscopy. There was significant difference among the control and any of the treated groups on the number of CK20 positive Merkel cells (p < 0.05) and that numbers were decreased at the sites where continuous mechanical stimulation was exerted. Degeneration of the cytoplasm mitochondria and nerve endings, and a decrease in both the number of neurosecretory granules and cytoplasmic processes were observed. Furthermore, the presence of nuclear chromatin aggregation and fragmentation was recognized. The continuous mechanical stimulation by the palatal plate affected the responses of Merkel cells and nerve endings, thus inducing a decrease in the number of Merkel cells. A portion of these changes was also associated with the expression of apoptosis.  相似文献   

12.
Fetal development of the unique papillary body and its localized peculiarities in the equine hoof are described based on the study of 51 fetuses, nine newborn foals, and five adult horses. The shape and dimensions of the dermal papillae and lamellae have a formative influence on the structure and physical quality of the corneous hoof capsule with its horn tubules and lamellae. The size and arrangement of these horn structures determine the mechanical quality of hoof horn. Proper horn quality is a prerequisite for the various functions of the hoof capsule, such as protecting the living dermis supporting the hoof capsule, shock absorption, and formation of the suspensory apparatus of the distal phalanx. Development of the segment-specific papillary body is initiated by the increasing mitotic activity of the epidermal cells invaginating the dermal surface, thus forming dermal microridges. These microridges are transformed into single dermal papillae, which are arranged in rows, or enlarged to become primary and secondary dermal lamellae. The formation of a segment-specific papillary body enables the increasing keratinization ratio in the hoof epidermis and the formation of the characteristic tubular and lamellar horn responsible for the special mechanical properties of hoof horn.  相似文献   

13.
A method has been developed for obtaining mixed primary cultures of dissociated epidermis enriched in Merkel cells. Merkel cells obtained from embryonic rat buccal pads were grown in serum-free medium and identified in vitro using a variety of histological and immunohistochemical markers. Quinacrine, a fluorescent amine, which has been used to identify Merkel cells in situ, labeled a morphologically distinct population of cells in vitro. Cells labeled with quinacrine had a large, phase bright nucleus with prominent nucleoli, surrounded by a phase dark perinuclear ring. Antibodies directed against neuron-specific enolase, another marker for Merkel cells in situ, and antibodies against a well-characterized neuroendocrine vesicle antigen also labeled this population of quinacrine fluorescent cells. Electron microscopic examination of our cultures indicated that cells containing characteristic features of Merkel cells including cytoplasmic dense-cored granules were present. A small but significant increase in the number of Merkel cells was observed over time in culture. Merkel cells supported the survival and outgrowth of both trigeminal ganglion sensory neurons and sympathetic neurons from the superior cervical ganglion in serum-free medium in the absence of exogenous nerve growth factor (NGF). Immunoblots probed with antibodies directed against NGF demonstrated that NGF was present in the medium taken from these cultures. NGF-like immunoreactivity colocalized to cells containing quinacrine fluorescence in situ and in vitro. Addition of antibodies directed against NGF to cocultures of Merkel cells and neurons decreased survival of sympathetic neurons by 90% and decreased survival of sensory neurons by 60%. These results suggest that Merkel cells are capable of providing trophic support for their normal complement of sensory neurons by producing NGF. Selective recognition of these targets was studied in vitro by characterizing the interactions between Merkel cells and growth cones from sensory or sympathetic neurons using both time-lapse videomicroscopy and standard morphometry of fixed cocultures. The majority of trigeminal ganglion sensory neurons (approximately 60%) extended growth cones onto clusters of Merkel cells. Neurites which contacted clusters of Merkel cells were significantly more highly branched than those growing on collagen. In contrast, the majority of sympathetic neurons (greater than 90%) failed to grow onto Merkel cells. Growth cones of sympathetic neurons often "collapsed" and retracted when contact was made with a cluster of Merkel cells. Fixation of Merkel cells with paraformaldehyde prior to coculture did not affect this difference between sensory and sympathetic neurite extension onto the Merkel cells. However, prior fixation of Merkel cells eradicated the apparent Merkel ce-induced branching of sensory neurites.(ABSTRACT TRUNCATED AT 400 WORDS)  相似文献   

14.
Gicerin/CD146 is a cell adhesion molecule, which belongs to the immunoglobulin (Ig) superfamily. We have reported that it has a homophilic binding activity, which participates in the neurite extension from embryonic neurons. To elucidate how gicerin is involved in the neurite extension mechanism, we employed PC12 cells, which expresses gicerin/CD146. PC12 cells extend longer neurites by nerve growth factor (NGF) on gicerin substrate than on without gicerin substrate, which indicates that gicerin participates in neurite extension by NGF. We also found that the expression of gicerin in PC12 cells is induced by NGF. Over-expression of gicerin also promotes neurite extension by gicerin-gicerin homophilic interaction. These findings suggested that increase of gicerin expression by NGF promotes the gicerin-gicerin homophilic interaction resulting in the neurite extension.  相似文献   

15.
Abstract: Genistein and other inhibitors of protein tyrosine kinases were examined for effects on neurite elongation and growth cone morphology in the rat PC12 pheochromocytoma cell line. Genistein increased the rate of neurite elongation in PC12 cells grown on a collagen/polylysine substratum after priming with nerve growth factor (NGF), but had no effect on undifferentiated cells. Steady-state levels of phosphotyrosine-modified proteins (105, 59, 52, and 46 kDa) were reduced in NGF-primed cells by genistein treatment. The target of genistein action did not appear to be the NGF receptor/ trk tyrosine kinase because the presence of NGF in cultures of NGF-primed cells was not necessary for genistein-stimulated neurite outgrowth. The tyrosine kinase inhibitors tyrphostin RG508964 and herbimycin A also increased the rate of neurite elongation in NGF-primed PC12 cells. Video-enhanced differential interference contrast microscopy revealed that growth cones of genistein-treated cells had less complex morphologies and were less dynamic than untreated cells, with short filopodia restricted to the leading edge, unlike untreated cells whose growth cones exhibited longer, more numerous filopodia and lamellipodia, which remodeled continuously. These results suggest that protein tyrosine kinase activity in PC12 cells negatively regulates neurite outgrowth and directly or indirectly affects growth cone morphology.  相似文献   

16.
Inactivation of Rho GTPases inhibited the neurite outgrowth of PC12 cells. The role of Cdc42 in neurite outgrowth was then studied by selective inhibition of Cdc42 signals. Overexpression of ACK42, Cdc42 binding domain of ACK-1, inhibited NGF-induced neurite outgrowth in PC12 cells. ACK42 also inhibited the neurite outgrowth of PC12 cells induced by constitutively activated mutant of Cdc42, but not Rac. These results suggest that Cdc42 plays an important role in mediating NGF-induced neurite outgrowth of PC12 cells. Inhibition of neurite outgrowth was also demonstrated using a cell permeable chimeric protein, penetratin-ACK42. A dominant negative mutant of Rac, RacN17 inhibited Cdc42-induced neurite outgrowth of PC12 cells suggesting that Rac acts downstream of Cdc42. Further studies, using primary-cultures of rat cerebellar granule neurons, showed that Cdc42 is also involved in the neurite outgrowth of cerebellar granule neurons. Both penetratin-ACK42 and Clostridium difficile toxin B, which inactivates all members of Rho GTPases strongly inhibited the neurite outgrowth of cerebellar granule neurons. These results show that Cdc42 plays a similar and essential role in the development of neurite outgrowth of PC12 cells and cerebellar granule neurons. These results provide evidence that Cdc42 produces signals that are essential for the neurite outgrowth of PC12 cells and cerebellar granule neurons. These authors contributed equally  相似文献   

17.
The aim of the present study was to investigate the responses of Merkel cells that are numerous in the palatine rugae, due to the continuous mechanical stimulation exerted by the palatal plate. Forty golden hamsters were used in this experiment. The palatal plate was made of adhesive resin and it was set on the palate of the animal. To exert a continuous pressure, a 0.8?mm elevation on the internal surface of the palatal plate was created at the middle portion of the fourth palatine ruga. Thereafter, the number of Merkel cells in the mucosa was calculated by immunohistochemical observation. Morphological changes of Merkel cells were examined by electron microscopy. There was significant difference among the control and any of the treated groups on the number of CK20 positive Merkel cells (p?<?0.05) and that numbers were decreased at the sites where continuous mechanical stimulation was exerted. Degeneration of the cytoplasm mitochondria and nerve endings, and a decrease in both the number of neurosecretory granules and cytoplasmic processes were observed. Furthermore, the presence of nuclear chromatin aggregation and fragmentation was recognized. The continuous mechanical stimulation by the palatal plate affected the responses of Merkel cells and nerve endings, thus inducing a decrease in the number of Merkel cells. A portion of these changes was also associated with the expression of apoptosis.  相似文献   

18.
Nijmegen breakage syndrome (NBS) is an autosomal recessive disorder, characterized by progressive microcephaly, growth retardation, immunodeficiency, and pre-disposition to tumor formation. To investigate the functions of the NBS gene product, NBS1, on neurons, PC12 cells overexpressing NBS1 and related mutants and primary cortical neuronal culture were used in the present study. Small interfering RNA (siRNA) was applied to repress the expression of endogenous Nbs1 in PC12 cells and primary cortical neurons. We demonstrated that overexpression of NBS1 increases cellular proliferation and decreases the apoptosis of PC12 cells in serum withdrawal and ionizing irradiation, through the activation of phosphatidylinositol 3-kinase (PI 3-kinase)/Akt pathway. Overexpression of NBS1 also decreases neurite elongation on PC12 cells under nerve growth factor stimulation. Transfection of NBS1-overexpressing PC12 cells with a dominant negative Akt mutant attenuates the neuroprotection and cellular proliferation effects of NBS1 while having no effect on neurite elongation. PC12 cells overexpressing NBS657del5 and NBS653 mutants, in which the major NBS1 protein in cells are truncated proteins, have decreased cellular proliferation, increased cell death, and decreased neurite elongation compared with those of control PC12 cells. Repression of Nbs1 by siRNA decreases the PI 3-kinase activity and Akt phosphorylation levels, and induces neurite elongation in PC12 cells even without nerve growth factor stimulation. Repression of Nbs1 by siRNA in primary cortical neurons also increased neurite elongation, but increased neuronal death. We conclude that NBS1 can regulate neuronal proliferation and neuroprotection via PI 3-kinase/Akt pathway while regulating neuronal differentiation in a different pathway. Excessive accumulation of truncated protein secondary to 657del5 mutation may be detrimental to neurons, leading to defective neuronal proliferation and differentiation.  相似文献   

19.
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.  相似文献   

20.
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.  相似文献   

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