Expression of alpha and beta adrenergic receptors in the pig uterus during inflammation.

Under physiological conditions, noradrenaline (NA) and adrenergic receptors (ARs) are implicated in the function of the uterus. The role of NA and the expression of ARs in the inflamed uterus is not fully understood. The aim of the present study was to determine the effect of inflammation on the levels of α1 (A, B, D)-, α2 (A, B, C)- and ß (1, 2, 3)-ARs mRNA and protein expression and the localization of these receptors in the porcine uterus. On Day 3 of the estrous cycle (Day 0 of the study), 50 ml of either saline (group SAL) or E. coli suspension (109 colony-forming units/ml, group E. coli) were injected into each uterine horn. In the control pigs (group CON), only laparotomy was performed. Eight days later, α1D-ARs mRNA (P < 0.001) and protein (P < 0.05) levels and α2A-ARs protein level (P < 0.05) were increased in the inflamed endometrium, while the α2C-ARs protein level (P < 0.001) was lowered, as compared to the SAL and CON groups. In the inflamed endometrium, ß2-ARs mRNA (P < 0.01) and protein (CON: P < 0.01, SAL: P < 0.001) expression was lower than in the other two groups, and ß1-ARs mRNA (P < 0.001) and protein (P < 0.01) expression was higher compared to the SAL group. After bacterial treatment, α2A- (P < 0.001) and α2B (P < 0.05) -ARs protein levels and ß2-ARs mRNA (CON: P < 0.01, SAL: P < 0.05) and protein (CON: P < 0.01, SAL: P < 0.05) expression in myometrium were found to be increased compared to both groups. In turn, in myometrium following E. coli infusion, the α2C-ARs protein level was lower (P < 0.01) than in the CON group. All studied receptors were present in the luminal and glandular epithelium, blood vessels and myometrial muscular cells of the gilt uteri in the E. coli, SAL and CON groups. The data show that inflammation changes the ARs expression in porcine uterus, suggesting their importance in the course/consequences of uterine inflammation. Those affected ARs may constitute a therapeutic target in an inflamed uterus.

Correction to: JNK-NQO1 axis drives TAp73-mediated tumor suppression upon oxidative and proteasomal stress.

Correction to: Cell Death Dis. 5, e1484 (2014); https://doi.org/10.1038/cddis.2014.408 ; published online 23 October 2014.

Long-term estradiol-17ß exposure decreases the cholinergic innervation pattern of the pig ovary.

Elevated levels of endogenous estrogens in the course of pathological states of ovaries, as well as xenoestrogens, may lead to hyperestrogenism. It has previously been demonstrated that long-term estradiol-17ß (E2) administration in adult gilts affected the population of sympathetic intraovarian nerve fibers. The aim of this study has been to determine the effect of long-term E2 exposure on the cholinergic innervation pattern of porcine ovaries. Intraovarian distribution and the density of nerve fibers immunoreactive (IR) to vesicular acetylocholine transporter (VAChT) and/or neuronal isoform of nitric oxide synthase (nNOS), vasoactive intestinal polypeptide (VIP), somatostatin (SOM) were determined. From day 4 of the first estrous cycle to day 20 of the second studied cycle, experimental gilts were intramuscularly injected with E2, while control gilts received corn oil. The ovaries were then collected and processed for double-labelling immunofluorescence. After E2 administration, the total number of fibers IR to VAChT, nNOS and VIP decreased significantly. The numbers of VAChT-, nNOS- and VIP-IR fibers within the ground plexus were significantly lower, while they were significantly higher around small or medium tertiary follicles. In the E2-affected ovaries, the numbers of nNOS- and VIP-IR fibers were significantly higher near secondary follicles and VAChT-IR in the vicinity of medullar blood vessels. In turn, around the latter structures there were significantly lowered populations of nNOS- and VIP-IR nerve fibers. These results suggest that the elevated E2 levels that occur during pathological states may affect the cholinergic innervation pattern of ovaries and their function(s).

Lipopolysaccharide and cytokines modulate leukotriene (LT)B4 and LTC4 production by porcine endometrial endothelial cells.

Uterine inflammatory response is mediated by inflammatory mediators including eicosanoids and cytokines produced by immune and endometrial cells. Interactions between lipopolysaccharide (LPS) and cytokines, and leukotrienes (LTs) in endothelium, important for the host defence during the inflammation, are unknown. We studied the effect of LPS, tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-4 and IL-10 on 5-lipooxygenase (5-LO), LTA4  hydrolase (LTAH) and LTC4 synthase (LTCS) mRNA and protein expression, LTB4 and LTC4 release from porcine endometrial endothelial cells, and cell viability. For 24 hr, cells were exposed to LPS (10 or 100 ng/ml of medium) and cytokines (each 1 or 10 ng/ml). 5-LO mRNA/protein expression augmented after incubation with larger doses of LPS, TNF-α, IL-4 and IL-10 and smaller dose of IL-1ß. Larger dose of TNF-α, smaller doses of LPS and IL-1ß and both doses of IL-10 increased LTAH mRNA/protein expression. LTAH protein content was up-regulated by larger dose of LPS, but it was reduced in response to both doses of IL-4. LTCS mRNA expression was elevated by larger doses of LPS, IL-4 and IL-10 or both doses of TNF-α and IL-1ß. LTCS protein level increased after treatment with both doses of IL-1ß, IL-4 and IL-10, smaller dose of LPS and larger dose of TNF-α. Both doses of LPS and larger doses of TNF-α and IL-10 increased LTB4 release. LPS, IL-1ß and IL-10 at smaller doses, or TNF-α and IL-4 at larger doses stimulated LTC4 release. Smaller doses of TNF-α and IL-1ß or both doses of IL-4 enhanced the cell viability. This work provides new insight on the participation of LPS, TNF-α, IL-1ß, IL-4 and IL-10 in LTB4 and LTC4 production/release from porcine endometrial endothelial cells, and the effect of above factors on these cells viability. The used cellular model gives the possibility to further establish the interactions between inflammatory mediators.

Porcine dorsal root ganglia ovarian neurons are affected by long lasting testosterone treatment.

We studied the effect of testosterone overdose on the number, distribution and chemical coding of ovarian neurons in the dorsal root ganglia (DRGs) in pigs. On day 3 of the estrous cycle, the ovaries of both the control and experimental gilts were injected with retrograde tracer Fast Blue. From day 4 of the estrous cycle to the expected day 20 of the second studied cycle, the experimental gilts were injected with testosterone, while the control gilts received oil. After the completion of the protocol the Th16-L5 DRGs were collected. Injections of testosterone increased the testosterone (~3.5 fold) and estradiol-17beta (~1.6 fold) levels in the peripheral blood, and reduced the following in the DRGs: the total number of the Fast Blue-positive perikarya, the population of perikarya in the L2-L4 ganglia, and the numbers of SP(+)/CGRP(+), SP(+)/PACAP(+), SP(+)/nNOS(+) and SP(-)/nNOS(+) perikarya. In the testosterone-injected gilts, the populations of SP(+)CGRP(-), small and large androgen receptors-expressing perikarya were increased. These results suggest that elevated androgen levels during pathological states may regulate the transmission of sensory modalities from the ovary to the spinal cord, and antidromic regulation of the ovarian functions.

JNK-NQO1 axis drives TAp73-mediated tumor suppression upon oxidative and proteasomal stress.

Hyperproliferating cancer cells produce energy mainly from aerobic glycolysis, which results in elevated ROS levels. Thus aggressive tumors often possess enhanced anti-oxidant capacity that impedes many current anti-cancer therapies. Additionally, in ROS-compromised cancer cells ubiquitin proteasome system (UPS) is often deregulated for timely removal of oxidized proteins, thus enabling cell survival. Taken that UPS maintains the turnover of factors controlling cell cycle and apoptosis--such as p53 or p73, it represents a promising target for pharmaceutical intervention. Enhancing oxidative insult in already ROS-compromised cancer cells appears as an attractive anti-tumor scenario. TAp73 is a bona fide tumor suppressor that drives the chemosensitivity of some cancers to cisplatin or γ-radiation. It is an important drug target in tumors where p53 is lost or mutated. Here we discovered a novel synergistic mechanism leading to potent p73 activation and cancer cell death by oxidative stress and inhibition of 20S proteasomes. Using a small-molecule inhibitor of 20S proteasome and ROS-inducer--withaferin A (WA), we found that WA-induced ROS activates JNK kinase and stabilizes phase II anti-oxidant response effector NF-E2-related transcription factor (NRF2). This results in activation of Nrf2 target--NQO1 (NADPH quinone oxidoreductase), and TAp73 protein stabilization. The observed effect was ablated by the ROS scavenger--NAC. Concurrently, stress-activated JNK phosphorylates TAp73 at multiple serine and threonine residues, which is crucial to ablate TAp73/MDM2 complex and to promote TAp73 transcriptional function and induction of robust apoptosis. Taken together our data demonstrate that ROS insult in combination with the inhibition of 20S proteasome and TAp73 activation endows synthetic lethality in cancer cells. Thus, our results may enable the establishment of a novel pharmacological strategy to exploit the enhanced sensitivity of tumors to elevated ROS and proteasomal stress to kill advanced tumors by pharmacological activation of TAp73 using molecules like WA.

Inhibitory effect of aluminium on the axonal transport of HRP microinjected into dorsal root ganglion neurons in vitro.

In the present study the function of axonal transport in individual neurons under aluminium intoxication was investigated experimentally in comparison with controls. We used the technique of microinjection of horseradish peroxidase (HRP) in dissociated dorsal root ganglia (DRG) neurons and neurons of explant cultures of DRG. Different exposure periods (1 and 6 hours as well as 6 and 10 days) to aluminium were analysed quantitatively. This analysis revealed an impaired anterograde transport of HRP already after a short aluminium intoxication period of only 1 hour in DRG cells in vitro, an effect that increased with a prolonged aluminium exposure for up to 10 days. Hence, functional alterations of the anterograde transport caused by aluminium could be detected even after short exposure periods. Furthermore, the effects of aluminium on anterograde transport mechanisms were reversible 8 days after removal of aluminium. To determine how aluminium affects the cytoskeleton, we performed immunohistochemistry and electron microscopy on cultured DRG neurons. Distinct morphological alterations of the cytoskeleton, especially the accumulation of phosphorylated neurofilaments, appeared after 6 days of aluminium exposure. Our results suggest that neurofilaments are indispensable to the functional integrity of the cytoskeleton and its ability to mediate microtubule-based axonal transport processes.

Taxol impairs anterograde axonal transport of microinjected horseradish peroxidase in dorsal root ganglia neurons in vitro.

We have investigated the effects of taxol on the axonal transport of horseradish peroxidase (HRP) in dorsal root ganglia (DRG) cells and their neuronal cytoskeleton. The former were analysed by microinjection of HRP into single DRG cells and the latter was studied by means of immunohistochemistry and cryo-electron microscopy. In cultured and untreated DRG cells, microinjected HRP was typically transported anterogradely several hundred micrometres along their neurites. Different exposure periods (1, 2 and 3 days) to taxol were analysed. The axonal transport of HRP in DRG cells was time-dependently impeded by taxol. After the drug had been washed out, a recovery of the axonal transport of HRP was observed and confirmed by quantitative analysis. Cryo-electron microscopy revealed an abnormal aggregation of axonal and cytoplasmic microtubules, associated with a decreased amount of cross-linking structures, in taxol-treated DRG cell cultures. After 3 days of taxol exposure, microtubule-associated proteins and Tau-protein were restricted to the cellular somata but the neurofilament network and tubulin-proteins seemed to be unaffected. Our results demonstrate, for the first time, an inhibition of anterograde axonal transport of HRP in single neurons by taxol. This effect is reversible and seems not to be caused by cellular damage, but is rather a consequence of an altered organisation of microtubules and/or microtubule-associated proteins.

Axonal and dendritic transport in Purkinje cells of cerebellar slice cultures studied by microinjection of horseradish peroxidase.

Axonal and dendritic transport in single Purkinje neurons of cerebellar slice cultures was quantified as single transport distances. Examination of the cells within a vital tissue was regarded as being an approach to the in situ condition. The Purkinje cells were organotypically integrated in the in vitro tissues and extended long axonal projections connecting synapses to the target neurons. The tracer horseradish peroxidase (HRP) was applied via microinjection to the somata of the Purkinje cells and the injected neurons were incubated thereafter for defined time-intervals. The tracer was transported anterogradely into the neuron processes. The measurements on both the axonal and the dendritic transport of microinjected HRP revealed continuous transportation with increasing times of postincubation. This transport was reduced by the use of microtubule-depolymerizing drugs. The axonal transport of the tracer was either retarded in colchicine-treated cells or continuously reduced for up to 50% in vinblastine-treated neurons. Thus, a correlation of axonal transport to the microtubules was demonstrated. The dendrites were filled with the tracer after 60 min of postincubation. Dendritic transport was reduced by the use of vinblastine, and not significantly by colchicine. The results strongly support the dependence of neuronal transport on microtubules as a component of the cytoskeleton.

Morphology of cryofixed myelin sheath.

The myelin sheath is formed by concentrically apposed membrane pairs and shows a regularly layered pattern of alternating light lines and dense lines. Observation of cryofixed myelin demonstrated that the structures represent aqueous spaces. All lamellae of the myelin sheath show globular aggregates of particles and these particles are corresponding with aggregates observed after detergent extraction of the myelin. Experimental fusion of myelin lamellae shows an intermixing of the globular particles or subunits. The interaction of these structural units in the bilayers may provide the stability of the myelin lamellae and their lamination.

Effects of different perfluorochemicals on dorsal root ganglion cells in vitro.

BACKGROUND: Investigation of the effects of different perfluorochemicals (PFC) on cultured dorsal root ganglion (DRG) cells. METHOD: DRG cell cultures from 9- to 11-day-old chicken embryos were exposed to emulsified perfluorodecalin (PFD; C10F18; 0.5%, 1% and 10%) or perfluorooctylbromide (PFO; C8F17Br; 0.5%, 1% and 10%). The cells were evaluated under phase-contrast optics after 30 h and 120 h for 0.5 and 1% and after 5 h for 10%. To study the integrity of neuronal cells, immunohistochemical labelling for neurofilaments (NF) and tubulin (TUB) was performed. RESULTS: Concentrations of 0.5% and 1% of PFD or PFO did not change immunohistochemical labelling of DRG cells. Co-cultured macrophages showed a foam cell response, presumably representing ingested PFC. At both concentrations PFD induced a weaker foam cell response than PFO. A concentration of 10% led to the death of DRG cells and macrophages within 5 h. CONCLUSION: PFC caused a dose-dependent damage of neuronal cells. Co-cultured macrophages developed a foam cell response similar to that observed in vivo after prolonged presence of PFC in the vitreous body. These observations indicate that PFD and PFO may not be suitable for long-term vitreous replacement in vitreoretinal surgery. However, the model is limited by several factors: (1) there are physiological differences between DRG cells and retinal ganglion cells; (2) in vivo retinal ganglion cells are protected by the overlying tissues; (3) the PFC used in tissue culture must be emulsified.

Pex17p of Saccharomyces cerevisiae is a novel peroxin and component of the peroxisomal protein translocation machinery.

The Saccharomyces cerevisiae pex17-1 mutant was isolated from a screen to identify mutants defective in peroxisome biogenesis. pex17-1 and pex17 null mutants fail to import matrix proteins into peroxisomes via both PTS1- and PTS2-dependent pathways. The PEX17 gene (formerly PAS9; Albertini, M., P. Rehling, R. Erdmann, W. Girzalsky, J.A.K.W. Kiel, M. Veenhuis, and W.-H Kunau. 1997. Cell. 89:83-92) encodes a polypeptide of 199 amino acids with one predicted membrane spanning region and two putative coiled-coil structures. However, localization studies demonstrate that Pex17p is a peripheral membrane protein located at the surface of peroxisomes. Particulate structures containing the peroxisomal integral membrane proteins Pex3p and Pex11p are evident in pex17 mutant cells, indicating the existence of peroxisomal remnants ("ghosts"). This finding suggests that pex17 null mutant cells are not impaired in peroxisomal membrane biogenesis. Two-hybrid studies showed that Pex17p directly binds to Pex14p, the recently proposed point of convergence for the two peroxisomal targeting signal (PTS)-dependent import pathways, and indirectly to Pex5p, the PTS1 receptor. The latter interaction requires Pex14p, indicating the potential of these three peroxins to form a trimeric complex. This conclusion is supported by immunoprecipitation experiments showing that Pex14p and Pex17p coprecipitate with both PTS receptors in the absence of Pex13p. From these and other studies we conclude that Pex17p, in addition to Pex13p and Pex14p, is the third identified component of the peroxisomal translocation machinery.

Human cornea and sclera studied by atomic force microscopy.

The purpose of this study was to investigate the ultrastructure of the extracellular matrix of human cornea and sclera by using the atomic force microscope (AFM). Specimens of human cornea (n=16) and sclera (n=10) were obtained from a cornea bank or from enucleated eyes (n=1; clinical and histopathological diagnosis: choroidal melanoma) and fixed in Karnovsky solution. The AFM resolved individual collagen fibrils in corneal and scleral tissue. Scleral collagen fibrils had a diameter ranging from 118.3 to 1268.0 nm and showed clear banding with a mean axial D-periodicity of 77.02 nm. The mean gap depth between the two overlaps was larger in the sclera than in the cornea. The diameter of corneal collagen fibrils ranged from 48.0 to 113.0 nm. In contrast to the sclera, the corneal collagen fibrils did not exhibit clear banding as their surface pattern. Closely attached fibrils with a beaded to globular structure were predominant in the cornea. The mean axial D-periodicity of the corneal collagen fibrils was 68.50 nm. In both tissues, the AFM resolved structures resembling cross-bridges between adjacent fibrils. The corneal collagen fibrils showed fibrillar properties that were different from those of the sclera, and that therefore might be essential for the spatial organization responsible for the optical quality of the cornea.

Cryo-electron microscopy of myelin treated with detergents.

After detergent extraction, myelin lamellae disaggregate into diverse globular fragments that correspond to the globular structures observed in normal cryofixed myelin. The interaction of these structural units in the bilayer is probably the morphological basis that provides the stability of the myelin lamellae and their lamination.

Axoplasmic transport of horseradish peroxidase in single neurons of the dorsal root ganglion studied in vitro by microinjection.

The dependence of anterograde axoplasmic transport on cytoskeletal components was investigated using microinjection of horseradish peroxidase (HRP) into the somata of chick dorsal root ganglion cells in vitro. Microinjected HRP was transported anterogradely in the neurites and their branches; this transport was disturbed by colchicine in a drug-dependent and time-dependent manner. Cytochalasin B, a drug that depolymerizes actin, did not inhibit the transport of HRP, despite the formation of local swellings in neurites. The microinjection of polyclonal antibodies directed against tubulin and monoclonal antibodies (mAbs) against 200-kDa neurofilaments disturbed the axoplasmic transport of co-injected HRP, which then exhibited an irregular and discontinuous distribution in the axonal branches. The transport of HRP became discontinuous after the injection of anti-tubulin antibodies and led to the formation of globular deposits of HRP. Polyclonal antibodies against actin and mAbs to 160-kDa and 68-kDa neurofilaments seemed to have no effect on the axoplasmic transport of co-injected HRP. Microinjection of antibodies against tubulin induced formation of perinuclear bundles consisting of cytoskeletal components. The transport of HRP thus appears to be regulated by an intact microtubular system and cross-linker components (200-kDa neurofilaments) of the cytoskeleton. Actin and most intermediate filament proteins do not seem to play an essential role in the transport of HRP.

[Results of therapy of anemia in pregnancy]. / Therapieergebnisse bei Schwangerschaftsanämie.

Among 106 pregnant women with anaemia a typical state of iron deficiency could be shown at only 36.8%. 22.5% of the patients had a decreased vitamin B12 level without any characteristic symptoms of a megaloblastic anaemia. Predominantly the grade of the anaemia was small. The mean value of hemoglobin lied at 7.1 +/- 0.59 mmol/l. The severity of the anaemia didn't show any connection to the vitamin B12 level or parameters of the iron metabolism. With a combined therapy of iron, folic acid and vitamin B12 an increase of the Hb-level could be noticed at only 44.3% of the patients. The haematological findings, taken before the therapy, as well as the therapy results show that an important part of anaemias in pregnancy is caused by a complex genesis as a result of immunological reactions in pregnancy.

Cryo-electron microscopy of nervous tissue. A review.

The present work attempts to demonstrate that cryofixation is a valuable method for the study of the nervous tissue. The use of the newly developed methods of cryofixation and freeze-etching without fixatives or cryoprotectants allows new exciting perspectives for the electron microscopical observation of cellular components, emphasizing their three-dimensional morphological structures. Significant contributions have been made on the fine structure of the cytoskeleton, cell membranes and cell organelles. The components of the cytoskeleton are distributed in different composition through the perikarya, dendrites and axon. The ubiquitous presence of the cytoskeleton suggests a crucial role in the functional activities of the neurons, especially in relation to the intracellular communication and to developmental and regeneration processes. Vitrified cellular membranes of myelin sheaths and rod outer segments have been observed in hydrated state by using cryofixation and cryotransfer techniques. These procedures allow new insights into the supramolecular structure and an approximation of morphological data to the present biophysical membrane model including a critical comparison with the current descriptions gained by conventional electron microscopy.

Cryo-electron microscopy of vitrified nerve myelin.

The ultrastructure of rat optic and trigeminal nerve myelin was studied using different cryotechniques. Replicas of rapid cryofixed and deep-etched material were compared with cryosections of chemically unfixed specimens and also of glutaraldehyde-fixed specimens. Hydrated cryosections were analysed in a cryotransfer device. The data reported here show discrepancies with the current descriptions of myelin structure based on osmium-fixed and resin-embedded material. The structures called the major line (as a fusion of the cytoplasmic surfaces of the glial cells) in conventional electron microscopy and the intraperiod line (as a fusion of the outer surfaces) are seen in the present material to represent actually aqueous spaces. The extracellular space (E-space) is most sensitive to chemical fixation and other preparation procedures, and probably also expands under pathological conditions. The virtual C-space (cytoplasmic space = major line) is more stable. The cytoplasmic surfaces are most probably joined by globular proteins (myelin basic protein). The most compact organization of myelin is seen in fresh, unfixed nerves. A continuous bilayer could not be observed and the bilayer membrane showed particulate subunits.

Chromatolysis of dorsal root ganglion cells studied by cryofixation.

Cytoskeletal alterations in the cytoplasm of chromatolytic neurons of the dorsal root ganglia were studied in chickens after transection of the sciatic nerves. These studies were carried out using cryofixation with a nitrogen-cooled propane jet. By this method, the morphological complexity of the cytoskeleton in normal perikarya and cell processes can be visualized. The cytoskeleton of the dorsal root ganglion cells (DRG) is composed of an intricate network of microtubules, neurofilaments and microfilaments. The membrane-bounded cell organelles, as well as the cell nucleus and the plasmalemma, are linked to the microtubules and neurofilaments by microfilaments (or cross-linkers). As a result of the transection of the axon, chromatolysis takes place, characterized by dislocation of cell organelles, an eccentric position of the nucleus and dispersion of the parallel cisternae of the rough endoplasmic reticulum throughout the cytoplasm. This characteristic phenomenon coincides with a regression of the neurocytoskeletal network. The neurofilaments and microtubules become shorter, and the microfilaments are replaced by strands of globular or granular material. The temporary regression of the microfilaments leads to a dispersion of the cell organelles. During the remodelling of the cytoskeletal structures, proliferation of the neurofilaments in the regenerating neurons may occasionally be observed. These results show that the cytoskeletal structures are responsible not only for the preservation of cell shape, but also for the maintenance of the normal distributional pattern (location and mobility) of the intracellular components.

Early structural changes in the axoplasmic cytoskeleton after axotomy studied by cryofixation.

Alterations in the cytoskeleton were studied in the axoplasm of neurites at the tips of proximal stumps of transected chicken sciatic nerves. The studies were carried out using cryofixation with a nitrogen-cooled propane jet. The most immediate effect is the almost complete disassembly of axoplasmic microtubules. This consequently causes the axonal transport of membrane-bounded organelles to cease and results in an accumulation of mitochondria and vesicles of the smooth endoplasmic reticulum. The neurofilament network is partially disorganized. Neurofilaments become shorter and fragmented, and are linked by a large number of anastomosed cross-linkers. The neurofilaments become newly aligned to the axis of the axoplasm and are of normal length 48-72 h after the transsection. At this stage the newly formed neurofilament bundles are in close proximity to the anastomosed cisternae and profiles of the smooth endoplasmic reticulum. The axonal sprouts always show a normally organized cytoskeletal network. These studies support the idea that the rapid remodelling of the neurofilament network is apparently a local event, not dependent on the slow transport of cytoskeletal materials to the tip of the proximal stump. The repair of the degraded cytoskeleton may be in accordance with the function of the endoplasmic reticulum as Ca2+-sequestering membrane system, which may be involved in restoring the physiological conditions of the axoplasm.