Peripheral nerve endoneurial microangiopathy and necrosis in rats with insulinoma

Peripheral nerve pathology related to chronic hyperinsulinemia and hypoglycemia has yet to be fully explored. Here we conducted a systematic quantitative analysis of morphological alterations in peripheral sensory and motor nerve fibers and endoneurial microvasculature in longstanding insulinoma-carrying rats (I-rats; n=12). Age-matched normal rats (n=6) served as controls. Over the 15-month observation period, two of I-rats developed paresis of the hind limbs when their blood glucose level fell below 1.7 mmol/l. These animals showed a massive myelinated fiber loss associated with active degeneration of residual myelinated fibers and multiple endoneurial microvascular occlusions at the sciatic nerve level. The rest of the non-paretic I-rats showed a decreased density of large myelinated fibers with axonal degeneration in the peroneal nerve and an increased density of small myelinated fibers with preserved morphology in the sural nerve. This was associated with endoneurial microangiopathic changes indicative of endoneurial ischemia/hypoxia in the sciatic and peroneal nerves, and an increase in endoneurial microvascular density in the sciatic and sural nerves. In conjunction with previous data, these findings suggest that the observed increase in endoneurial microvascular density may be a compensatory response to endoneurial ischemia/hypoxia induced by chronic hyperinsulinemia in I-rats without paresis. In conclusion, the present study showed characteristic morphological alterations in peripheral sensory and motor nerve fibers associated with microangiopathy indicative of endoneurial ischemia/hypoxia in the sciatic and peroneal nerves, and provides the first evidence for the occurrence of endoneurial necrosis in the sciatic nerve, to which the hind limb paresis can be ascribed in I-rats.     

Endoneurial microenvironment and acute nerve crush injury in the rat sciatic nerve

In severe peripheral nerve ischemia in the rat, serial nerve blood flow (NBF) measurements have identified evidence of 'no reflow', a mechanism of continued fiber damage during reperfusion. It has been postulated that 'no reflow' also occurs in nerve compression due to direct mechanical or ischemic (if compression is prolonged) injury of microvessels, resulting in continuing nerve fiber damage. To address this question, we measured endoneurial blood flow (NBF), oxygen tension and pH at the site of an acute nerve crush injury. In further sets of experiments, NBF and endoneurial oxygen tension were examined before and after prolonged epochs of crush. NBF and MR (microvascular resistance) were not appreciably different than values obtained in control animals without intervening brief nerve crush. NBF was slightly higher and MR slightly lower 2 h after injury, but the difference was not statistically significant. No evidence of significant endoneurial hypoxia or acidosis was observed. Similarly, after more prolonged crush there was no significant oligemia or hypoxia. The studies provide no evidence that 'no reflow' occurs in crush injury even if the injury is maintained for a period of time known to induce 'no reflow' with severe ischemia. We suggest that nerve damage in crush, and possibly compression, more likely arises from direct mechanical injury of fibers.     

Quantitative and qualitative study of sural nerve biopsies in Krabbe's disease

Summary Sural nerve biopsies from two infants with Krabbe's disease were examined morphologically, and quantitatively analyzed to obtain the density and size distribution of myelinated and unmyelinated nerve fibers as well as the variation in internodal lengths along the course of teased nerve fibers. When compared with age-matched control material, these sural nerves revealed a 50% reduction in density of myelinated fibers with a relative preponderance of small myelinated fibers. It was estimated that each nerve contained about 15% of the large (7–10) nerve fiber population found in control nerve. No reduction in the density of unmyelinated fibers was detected.Studies of teased preparations revealed an intermittent shortening of myelin internodes with resulting increased disparity of internodal lengths along individual fibers. These changes were indicative of widespread and severe demyelination and remyelination involving peripheral myelinated nerve fibers of all sizes in Krabbe's disease. No evidence of axonal or Wallerian degeneration was detected.Lipid accumulations were seen within endoneurial macrophage and within Schwann cells of myelinated fibers in both sural nerves and in the sympathetic chain removed at autopsy in case 2. The Schwann cell deposits were osmiophilic and appeared as focal collections of paranodal granules along myclinated fibers of teased preparations.Supported by U. S. P. H. S. Grants No. NB 08620 and NS 08054, and grants from the Allen P. and Josephine B. Green Foundation, Mexico, Missouri.     

Abnormalities of peripheral nerve in patients with human immunodeficiency virus infection

The purpose of this study was to assess systematically morphology of peripheral nerves from patients with human immunodeficiency virus infection (acquired immunodeficiency syndrome [AIDS] and AIDS-related complex) examined at autopsy. Sural nerve specimens were taken from 25 patients (mean age 44 years) and evaluated by routine procedures used in our laboratory. In 13 cases no detectable abnormality was seen. Twelve patients (48%) showed loss of myelinated fibers with disproportionately greater loss of large myelinated fibers. Three of these patients showed severe myelinated fiber loss; 2 had no documented symptoms and no other known predisposing factors for a peripheral neuropathy. Changes suggestive of wallerian degeneration were occasionally seen, as were epineurial and endoneurial inflammatory infiltrates. Segmental demyelination was not identified in any nerve examined. Electron microscopy revealed thickened basement membranes around small blood vessels, Schwann cells, and fibroblasts. Peripheral nerve abnormalities in patients with AIDS or ARC are frequent and their pathogenesis remains unclear.     

Changes in motor nerve terminals following proximal constriction by a ligature

To elucidate the effect of proximal constriction on motor nerve terminals, silk ligations were placed around the tibial nerve in the thigh of rabbits. The ligatures were tight enough to cause Wallerian degeneration in most of the large myelinated fibers; we studied those which remained unaffected. A week after operation, 9 animals showed a fall in amplitude of medial plantar muscle action potential to less than 30% of the pre-operative value on tibial nerve stimulation at the ankle. They were killed after keeping the constriction from 10 to 100 days, and the medial plantar muscles were removed for histological studies on the motor terminals of the medial plantar nerve. AChE-silver staining showed many nerve endings without terminal axons, and "junctional" terminals showing preservation of the continuity proximal to complete degeneration from 10 days to the 40 days after ligation. A few terminal and nodal sproutings were found 10 days after ligation. Transverse sections of the intramuscular portion of the medial plantar nerve showed a decrease in number of the large myelinated fibers. While the ratio of axonal caliber/external diameter of large myelinated fibers (g-ratio) was reduced, g-ratio of small myelinated fibers were varied but as high as that in normal controls from 40 days after ligation. These results indicate distal axonal degeneration (dying back) of the terminal fibers besides the Wallerian degeneration at the level of the ligature and inhibited distal sproutings, which are probably caused by a local disturbance of axonal transport resulting from proximal constriction.     

An experimental painful peripheral neuropathy due to nerve constriction. I. Axonal pathology in the sciatic nerve.

A constriction injury to the sciatic nerve of the rat produces a painful peripheral neuropathy that is similar to the conditions seen in man. The pathology of the sciatic nerve in these animals was examined at 10 days postinjury, when the abnormal pain sensations are near maximal severity. The nerves were examined with (1) complete series of silver-stained longitudinal sections of pieces of the nerve (3 cm or more) that contained the constriction injury in the center, (2) toluidine blue-stained semithin sections taken at least 1 cm proximal and 1 cm distal to the constriction, and (3) EM sections taken adjacent to those stained with toluidine blue. One centimeter or more proximal to the constriction, both myelinated and unmyelinated axons were all normal. Nearer to the constriction, extensive degeneration of myelinated axons became increasingly common, as did signs of endoneurial edema. Distal to the constriction, the nerve was uniformly edematous and full of myelinic degeneration. There was a profound loss of large myelinated axons and a distinctly less severe loss of small myelinated and unmyelinated axons. These observations show that at 10 days postinjury the constriction produces a partial and differential deafferentation of the sciatic nerve's territory. The absence of degeneration in the nerve 1 cm proximal to the constriction indicates the survival of the primary afferent neurons whose axons are interrupted.     

Morphological study of the aging human peripheral nerve

The superficial peroneal nerve from 46 elderly patients was studied by light and electron microscopy. Like most elderly people, these patients displayed evidence of varying degrees of peripheral nervous system dysfunction, but they were not suffering from any disease known to alter the peripheral nerve. A quantitative study showed that myelinated fiber loss predominated in the large diameter group. In the majority of cases, clusters of regenerating myelinated fibers were numerous. On the other hand, Wallerian-like degeneration aspects, segmental demyelination figures and axonal organelle accumulation were relatively rare. Unmyelinated fibers were also notably damaged in most cases. There was moderate reduplication of endoneurial vascular basement membrane.     

Cellular pathology of the nerve microenvironment in galactose intoxication

The effect of chronic hyperglycemia and polyol pathway activation on the Schwann cell has not been resolved although injury to this cell has long been suspected in diabetic neuropathy. Hyperglycemia, resulting from galactose intoxication of four months duration, induces dose-dependent accumulations of endoneurial fluid sodium and chloride that are linked to polyol pathway activity and associated with dose-dependent increases in sciatic nerve water content, endoneurial fluid pressure and (Na+, K+)-ATPase activity. In order to understand the impact of these changes on the nerve microenvironment, cellular elements of the endoneurium were quantitatively and qualitatively assessed in rats receiving 0%, 10%, 20% or 40% galactose diets. After four months of galactose intoxication, dose-dependent changes in the size distribution of myelinated nerve fibers were apparent. A shift in size-frequency histograms of galactose-intoxicated animals towards smaller fibers was accompanied by a decrease in axon diameter and the volume fraction ratio of axon to myelinated nerve fibers. In the sciatic nerve of all 40% galactose-fed rats examined by electron microscopy, Schwann cells of myelinated fibers showed both reactive and degenerative changes. Demyelination was preceded by splitting at the intraperiod line. Remyelination was identified by axons with disproportionately thin myelin sheaths. Axonal dystrophy and degeneration were infrequently seen, but there was axonal regeneration. Dose-dependent increases in mast cell number were observed with degranulation apparent in rats receiving 20% and 40% galactose. Endothelial cell number and basal lamina thickness were increased in the endoneurial vessels of galactose-intoxicated rats. Increased cytoplasmic area and degenerative changes in pericytes were also noted. These observations indicate that significant morphologic changes accompany the hyperosmotic imbalance resulting from galactose intoxication of four months duration. Schwann cell injury and demyelination are present in a disorder linked to polyol metabolism since aldose reductase, the anabolic enzyme of the polyol pathway, is localized to this myelin-forming cell.     

Perineurium contributes to axonal damage in acute inflammatory demyelinating polyneuropathy

OBJECTIVE: To assess if axonal damage in severe acute inflammatory demyelinating polyneuropathy (AIDP) correlates with the appearance of epiperineurium in nerve trunks. BACKGROUND: Increase of endoneurial fluid pressure in nerve trunks possessing epiperineurium may be an important mechanism of axonal damage in AIDP. METHODS: A 79-year-old man had a 2-day history of acroparesthesias and ascending paralysis culminating in quadriplegia, bilateral facial palsy, and mechanical ventilation. Five intravenous immunoglobulin cycles were given without response. He died on day 60. Electrophysiologic studies (days 4, 17, and 50) initially showed normal nerve conduction velocities with further slowing, progressive attenuation of compound muscle action potentials, and profuse denervation. The authors studied the preforaminal anterior and posterior L3 and L5 spinal roots, third and fifth lumbar nerves and their branches, and femoral and sural nerves. RESULTS: Density of myelinated fibers was preserved in L5 ventral and dorsal roots and reduced in sural nerve. Mild de-remyelination was observed in lumbar roots. In both lumbar nerves and their branches, there were extensive de-remyelination and centrofascicular or wedge-shaped areas with marked loss of large myelinated fibers. Axonal degeneration was the predominant lesion in sural nerve. CONCLUSION: The presence of epiperineurium correlates with a drastic change of pathology with superimposed ischemic lesions and distally accentuated axonal loss, suggesting that endoneurial fluid pressure increase could cause axonal damage in AIDP.     

Dysmaturation neuromyopathy: correlation with minimal neuropathy in sural nerve biopsies

Dysmaturation (neuro)myopathy without specific histochemical or cytoarchitectural characteristics accounts for many cases of hypotonia in infancy. We obtained a maturation profile for type I and type II fibers from birth to 6 years of age, from which a classification of fiber dysmaturation based on fiber-type hypotrophy and coefficient of fiber variation is presented. We analyzed the morphometric, ultrastructural, and single fiber teasing findings in the sural nerve of ten infants with hypotonia and dysmaturation myopathy based on the above classification. Data on endoneurial area, myelinated fiber density, proportion of large (greater than 6 micron) myelinated fibers, unmyelinated fiber density, regression analyses of myelin area:axon area, and mean myelin thickness were developed. Abnormalities in large myelinated fiber density (3 cases), disturbances in myelination index (5), and single fiber teasing abnormalities in internode length (1) were found. Ultrastructural abnormalities were observed and classified as group 0 changes similar to the five control nerves; group 1, in which rare fibers showed degeneration or basement membrane duplication; and group 2, which was characterized by multiple abnormalities including degenerate nuclei, amyelinate fibers, degeneration/regeneration, excessive basal lamina reduplication, and hypomyelination. This study provides evidence that minimal neuropathic abnormalities are present in the sural nerve of infants carrying a diagnosis of dysmaturation neuromyopathy.     

Peripheral nerve pathology in rats with streptozotocin-induced insulinoma

Peripheral nerve structure was systematically examined in rats with insulinoma induced by streptozotocin (STZ). Normal Wistar rats, aged 3 months (n = 10), were treated with intravenous injections of STZ (20 mg/ kg) and housed in plastic cages with free access to water and chow until 24 months of age. Three rats with insulinoma survived and were examined pathologically. Age-matched normal Wistar rats (n = 6) were used for comparison. The insulinoma-bearing rats showed a marked increase in body weight and decrease in blood glucose. In a teased nerve fiber study of the sciatic nerve, the percentage of abnormal fibers undergoing axonal degeneration and de- and remyelination in age-matched normal control rats was 3.9 ± 2.5% (means ± SD), whereas in the three insulinoma-bearing rats 49%, 50%, and 24%, respectively, of the fibers showed such changes. Regenerating fibers were also numerous in each insulinoma-bearing rat (36%, 42% and 27%, respectively). Morphometric analysis revealed smaller mean myelinated fiber and axonal areas in all the nerves examined (sciatic, tibial and sural) in insulinoma-bearing rats as compared to those in age-matched normal rats. Fiber area frequency histograms showed a decrease in large myelinated fibers and an increase in small regenerated fibers in insulinoma-bearing rats. Ultrastructurally, endoneurial microvessels exhibited a narrowed vascular lumen with swollen endothelial cells and vacuolar degeneration of pericytes, suggesting an involvement of vascular changes in the neuropathic development. The present study demonstrated marked structural changes in both motor and sensory peripheral nerves of rats bearing experimentally induced insulinoma. We consider that axonal degeneration, regeneration and demyelination constitute the main pathology in the peripheral nerves of insulinoma-bearing rats, although no particular difference in severity of the lesions between sensory and motor and between proximal and distal nerves was apparent. Received: 24 October 1995 / Revised, accepted: 27 December 1995     

Marklose nervenfasern im Senium und im Spätstadium der Thalidomid-Polyneuropathie: quantitativ-elektronenmikroskopische Untersuchungen

Summary Sural nerve biopsies of four patients, aged 54–76 years, with a predominantly sensory type of neuropathy following high dosages of thalidomide were examined by light and electron microscopy. The present study includes a qualitative and quantitative evaluation of unmyelinated nerve fibers. Despite severe neuropathy, increased numbers of small unmyelinated axons per endoncurial area were noted in all patients. This numerical increase appeared to be independent of aging, since it was not seen in two senile controls, studied at the age of 83 and 88 years. The increase in the endoneurial density of unmyelinated axons, especially of small sized fibers, is likely to be related to regeneration following degeneration of unmyelinated axons although endoneurial shrinkage secondary to loss of large myelinated fibers could have caused an additional increase in the number of axons per endoneurial area.Axonal sprouting, despite degeneration of large numbers of myelinated and unmyelinated fibers, appears to be consistent with some of the characteristic clinical features of thalidomide neuropathy such as paresthesias, hyperesthesia for pain and temperature, and disturbances of autonomic functions. On the other hand, a variable number of empty Schwann cells (bands of Büngner) and pockets at the surface of many Schwann cells noted in the four patients with neuropathy were also seen in both senile controls with no signs of neuropathy. Thus, it is obvious that pockets and empty Schwann cells may be related to aging or other causes of slow axonal wasting with Schwann cell proliferation and are not necessarily associated with clinically manifest neuropathy.

Mit Unterstützung der Deutschen Forschungsgemeinschaft, Bonn-Bad Godesberg (Schr 195/1)     

Ultrastructural study of normal and degenerating nerve fibers in the protocerebral tract of the crab Ucides cordatus

Wallerian degeneration is a very well described phenomenon in the vertebrate nervous system. In arthropods, and especially in crustaceans, nerve fiber degeneration has not been described extensively. In addition, literature shows that the events do not follow the same patterns as in vertebrates. In this study we report, by qualitative and quantitative ultrastructural analyses, the features and time course of the protocerebral tract degeneration following extirpation of the optic stalk. No remarkable changes were observed seven days after lesion. After 28 days the protocerebral tracts presented apparently preserved small and large diameter axons and some degenerating medium axons, with irregular contours and empty-looking aspect of the axoplasm. Forty days after the ablation of the optic stalks, both small (type I) and medium (type II and III) axons revealed signs of partial or total degeneration, but large nerve fibers (type IV) were still intact. After 45 days, the tract showed signs of advanced stage of degeneration and, apart from large axons, normal-looking fibers were almost absent. At these 3 last time points, degenerating axons displayed different electron densities and aspects, probably correlating to different onset times of the process. In addition, cells with granules in their cytoplasm, possibly hemocytes, were quite distinct, especially at 40 and 45 days after axotomy. These cells might share with glial cells the function of phagocytosis of cellular debris during the protocerebral tract degeneration. Quantitative analysis showed that the number of degenerating fibers increased significantly from 28 to 40 days after lesion, whereas the number of normal fibers decreased accordingly. Measurements of cross-sectional areas of normal and degenerating axons showed that types II and III (medium) start to degenerate before type I (small). Type IV (large) axons do not degenerate, even after 40 days. Therefore, we can conclude that degeneration in these afferent fibers starts late after axotomy, but proceeds at a faster rate afterwards until the complete degeneration of small and medium axons.     

Licht- und elektronenmikroskopische sowie cytometrische Untersuchungen an peripheren Nerven beim Morbus Tangier

Summary A sural nerve biopsy was performed in a 55-year-old male patient with Tangier disease (familiallipoprotein deficiency). Light-microscopy showed an increase in the endoneural connective tissue and a loss of nerve fibers indicating a chronic peripheral neuropathy. Electron-microscopy revealed an accumulation of lipid droplets within Schwann cells of myelinated and unmyelinated nerve fibers. When compared with age-matched controls the myelinated fiber density was reduced with a relative preponderance of small myelinated fibers. In addition, distributional cytometric studies of nerve vibers in relation to the perineurium and endoneurial capillaries showed: Contrary to 4.6–7.5 m thick nerve fibers, which accumulated in the center of the nerve fascicle, small (0.5–4.5 m) and large (7.6–10.0 m) fibers lay nearby the perineurium. The measured increase in small myelinated nerve fibers around endoneurial capillaries may be explained as a sign of regeneration.

     

Photochemically-induced ischemic injury of the rat sciatic nerve: a light- and electron microscopic study

Lesion of presumably ischemic origin of the rat sciatic nerve was induced photochemically by laser irradiation combined with systemic administration of a photosensitizing organic dye, erythrosin B. We have studied the pathologic features of the nerve after the photochemical insult with light- and electronmicroscopy and related them to behavioral signs of neuropathic pain. At the irradiated nerve site, occlusion of blood vessels was seen and the vessels were packed with aggregated thrombocytes, fibrins and deformed erythrocytes, supporting the notion that photochemical reaction caused intraneural ischemia. The degree of the nerve injury at the center of irradiation was related to the duration of the laser exposure. Brief irradiation (30 seconds) only caused identifiable injury to myelinated fibers, whereas longer irradiation (2 minutes) caused greater injury to myelinated and unmyelinated fibers, characterized by extensive axonal degeneration and demyelination. The rats irradiated for 2 minutes, but not 30 seconds, exhibited neuropathic pain-like behaviors, expressed as mechanical and cold allodynia. The nerve injury was most severe 7 days after ischemia and regeneration of both myelinated and unmyelinated fibers was observed 3 months later. The nerve caudal to the irradiation exhibited Wallerian degeneration 7 days after the insult, whereas at 10 mm proximal to the irradiation the nerve was largely normal. It is thus concluded that photochemically induced intraneural ischemia caused injury to both myelinated and unmyelinated fibers, with myelinated fibers being more susceptible. However, the development of neuropathic pain-like behaviors may require injury to the unmyelinated fibers.     

Efficacy of limb cooling on the salvage of peripheral nerve from ischemic fiber degeneration

Since peripheral nerve has a large ischemic safety factor, hypothermia, by reducing metabolic demands, it is potentially an efficacious technique to rescue nerve from ischemic fiber degeneration (IFD). We therefore evaluated the influence of temperature on the severity of IFD resulting from a standard ischemic stress. Ischemia to the left sciatic nerve in the rat was produced by embolization of 2 × 10⁶ microspheres (14 μm) into its supplying arteries. The limb was embolized at three temperatures, 37°C, 32°C, and 28°C and was maintained at each temperature for an additional 4 h. End points, evaluated 7 days after embolization, for the embolized limb were: (1) behavioral scores, 0–11 in increasing limb function; (2) compound nerve action potential of sciatic-tibial nerve; (3) sciatic nerve blood flow (NBF, in mL/100 g/min); and (4) histologic grade, expressed as percentage of fibers undergoing IFD (0 = < 5% 1 = 5–25% 2 = 26–50% 3 = 51–75% 4 = >75%). NBF was reduced in all groups, varying with temperature, and all indices of nerve structure and function were significantly improved with hypothermia. We conclude that hypothermia, easily achievable in a limb nerve, is highly efficacious in the rescue of nerve from IFD. These findings are of clinical relevance. © 1996 John Wiley & Sons, Inc.     

Sural nerve biopsy findings in leprosy: a qualitative and quantitative light and electron microscope study in 4 treated cases of the lepromatous spectrum

Reports on biopsy findings in multifascicular nerves in lepromatous leprosy (LL) are rare and detailed morphometrical data are not available. In a case of early LL with normal electrodiagnostic findings in sural nerve, the present study revealed marked segmental de- and remyelination concomitant with the sequelae of considerable Wallerian degeneration of preferentially small myelinated fibers (MF) in spite of a normal number/nerve and density/mm2. Segmental de- and remyelination of several consecutive internodes in teased fibers suggests continuous bacterial spread via Schwann cells. In 2 more advanced LL-cases, nervous parenchyme was severely reduced, in a borderline lepromatous (BL) case obviously in part caused by cell infiltrates and granulomata. Distinct fascicle differences in MF-involvement were demonstrated by coefficients of variation of MF/mm2 and teased fiber preparations in LL, consistent with the hypothesis of initial focal spread of bacteria. Numbers and densities of endoneurial vessels were increased only in the later stages of LL. Enlargement of endoneurial area, due to different factors, was encountered except for the most severe LL-case with extensive endoneurial collagenization. Morphometric results were compared with those of other neuropathies. Intact and degenerating bacteria mostly in phagosomes of Schwann cells associated with unmyelinated axons and in macrophages were seen only in the early LL- and the BL-case. Sparse mononuclear cell infiltrates and small focal necrosis, present even in LL-cases, underline the complex pathogenesis of nerve fiber involvement.     

Blood-nerve barrier in IgM paraproteinemic neuropathy: a clinicopathologic assessment

We report the pathologic findings in a patient with sensorimotor neuropathy associated with Waldenström’s macroglobulinemia, particularly in relation to blood-nerve barrier defects. The monoclonal IgM was of κ type and possessed anti-HNK-1 activity. A sural nerve biopsy specimen revealed severe loss of myelinated and unmyelinated nerve fibers and gaps between adjacent endothelial cells of small endoneurial vessels. Postmortem findings 3 years later included severe loss of myelinated nerve fibers and diffuse infiltration by lymphoplasmacytic B cells throughout the peripheral nervous system, sparing the central nervous system. Findings in this case suggest an immune attack against endoneurial endothelial cells with permeation of IgM into peripheral nerve tissue.     

Reperfusion nerve injury: pathology due to reflow and prolonged ischaemia

Ischaemia plays an important role in the development of pathological changes in various neuropathies. Nerve pathology in acute ischaemic injury has been delineated longitudinally in peripheral nerve and reperfusion injury could amplify ischaemic pathology. We examined ischaemia/reperfusion-induced pathological changes along the length of sciatic, tibial and peroneal nerves from pelvic to ankle levels. Pathological features were correlated with the degree of postischaemic nerve blood flow (NBF) restoration, measured by a laser Doppler flowmeter, and the blood-nerve barrier function by a horseradish peroxidase (HRP) technique. Major arteries which supply rat hindlimb were occluded for 3, 5, or 7 hours, and reperfusion was accomplished by the removal of vascular clips. Nerve pathology was assessed after 12, 24 and 48 hours, and 5 and 7 days of reperfusion. Pathological alterations at the thigh level included vascular swelling, endoneurial and intramyelinic oedema, demyelination, thrombosis, and red blood cell (RBC) extravasation. A paucity of axonal degeneration was also characteristic at this level. By contrast, the distal nerve from knee to ankle showed evidence of extensive axonal changes, occluded vessels, and panfascicular nerve fibre and vascular degeneration. Postischaemic NBF reflow was confirmed at the thigh level immediately, 24 and 48 hours after reperfusion, whereas NBF restoration at the knee to calf level was less than preischaemic values. HRP leakage was found in both proximal and distal nerve segments. In conclusion, we demonstrated different types of structural changes along the length of ischaemic/reperfused rat sciatic nerves. The present study suggests that pathological abnormalities at the thigh level are most likely due to reoxygenation of ischaemia-inflicted endothelial cells, and distal morphological changes could be induced by prolonged ischaemia resulting from occluded vessels.     

Biophysical and pathological effects of cryogenic nerve lesion

Changes in endoneurial fluid pressure (EFP) and morphology were studied in rat sciatic nerves frozen for 60 seconds with a cryoprobe designed for human cryoanalgesia. The onset of increased EFP was rapid, and a peak of 23 cm H₂O was reached within 90 minutes after injury. EFP levels returned to normal 32 days after freezing. The peak value represents the highest EFP yet recorded in an experimental neuropathy. Microscopic examination revealed severe vascular injury as the probable mechanism of edema, with leakage of horseradish peroxidase tracer at the site of injury and diapedesis of polymorphonuclear cells through vessel walls. Wallerian degeneration was also observed in segments of nerve distal to the site of injury. Analysis of EFP data revealed a biphasic pattern of endoneurial edema: initial marked pressure elevation subsides within hours but is followed by a second peak several days later. We interpret this to suggest superposition of two separate pathological processes following cold injury. At first, extensive vascular damage permits plasma and cellular extravasation, which rapidly increases EFP. Subsequently, nerve fibers undergo wallerian degeneration, a process associated with elevated EFP, which is maximal 6 days after injury.