Prior collateral sprouting enhances elongation rate of sensory axons regenerating through acellular distal segment of a crushed peripheral nerve

Regenerating axons in crushed peripheral nerves grow through their distal nerve segments even in the absence of Schwann cell support, but their elongation rate is reduced by 30%. We examined whether prior exposure of sensory neurons to trophic factors achieved either by collateral sprouting or regeneration after conditioning lesion could enhance subsequent regeneration of their axons after crush, and compensate for loss of cell support. Collateral sprouting of the peroneal cutaneous sensory axons in the rat was evoked by transection of adjacent peripheral nerves in the hind leg. The segment of the peroneal nerve distal to the crush was made acellular by repeated freezing. Sensory axon elongation rate during regeneration was measured by the nerve pinch test. Prior axonal sprouting for two weeks increased the elongation rate of sensory axons through the acellular distal nerve segment back to normal value observed in control crushed nerves. The number of axons in the acellular distal segment at a fixed distance from the crush site was about 50% greater in sprouting than in control non-sprouting nerves. However, prior sprouting caused no further increase of axon elongation rate in control crushed nerves. Prior collateral sprouting, therefore, could in some respect compensate for loss of cell support in the distal nerve segment after crush lesion. This suggests that loss of cell-produced trophic factors is probably responsible for slower elongation rate through the acellular distal nerve segment. Surprisingly, prior conditioning lesion caused no enhancement of elongation rate of the sensory axons regenerating in the absence of cell support.     

Schwann cell dependence of regenerating rat sensory neurons is inversely related to the quality of axon growth substratum

It is still controversial to what extent elongation of regenerating sensory axons depends on proliferating Schwann cells (SCs) in an injured peripheral nerve. We hypothesized that such regeneration was independent of SC support early after nerve injury, but later became SC-dependent. The sural nerve in rats was crushed, and freezing destroyed cells but not their basal laminae (BL) in the distal nerve segment. Sensory axon elongation was assessed by the nerve pinch test and their abundance was examined immunohistochemically. Sensory axons regenerated fairly rapidly during the first week even if SC migration was prevented. Thereafter, they ceased to elongate and withdrew until their terminals contacted the SCs migrating from the proximal nerve segment. Intrinsic neuronal capacity for growth without cell support, however, had not been lost. Rather, progressive degradation of the former SC BL and loss of laminin in the acellular segment arrested axon growth. Further elongation occurred only when SC migration was possible, corroborating our hypothesis. Sensory neurons continued to elongate and maintain their axons in spite of deteriorating growth substratum if, prior to injury the axons had been allowed to sprout into the denervated skin. Previous sprouting exposed the sensory neurons to high levels of NGF.     

Neurite-promoting influences of proliferating Schwann cells and target-tissues are not prerequisite for rapid axonal elongation after nerve crush

An important role in peripheral nerve regeneration has been ascribed to humoral trophic and tropic agents arising from the nonneuronal cells in the distal nerve stump and the denervated targets. In order to estimate their contribution to axonal elongation after crush injury to the rat sciatic nerve, an in vivo model was designed in which local cellular and target-derived influences were eliminated by 1) freeze-thawing of a long nerve segment distal to the crush site and 2) cutting the nerve far distally to the crush site, but within the frozen-thawed segment, and deflecting the frozen-thawed nerve stump in the opposite direction from its natural course. The sensory and motor axon elongation rate was estimated from the results of the nerve pinch test and choline acetyltransferase distribution along the nerve segment distal to the crush. The elongation rate of regenerating axons in deflected nerve segments, either non-treated or frozen-thawed, was close in magnitude to that obtained when target-derived influences were not eliminated. Neurotropism of axonal targets is therefore of little importance for axon elongation after nerve crush. In the absence of Schwann cells along the axonal path in frozen-thawed nerve segments, the elongation rate of both sensory and motor axons declined by about 40%. This implies that interactions between viable Schwann cells and growth cones of regenerating axons are not prerequisite for rapid axon elongation when Schwann cell basal lamina constitutes the growth substratum. Nevertheless, Schwann cells in Bungner bands possibly enhance the axon elongation rate by humoral or cell surface-mediated mechanisms.     

Prior Collateral Sprouting of Sensory Axons Delays Recovery of Pain Sensitivity after Subsequent Nerve Crush

Regeneration of motor axons is enhanced if they have sprouted prior to nerve injury. We examined whether sensory axon regeneration and recovery of pain response was affected by previous collateral sprouting. In the experimental group of rats, the right saphenous, tibial, and sural nerves were transected and ligated. The peroneal nerve was left to sprout into the adjacent denervated skin. Two months later, the axons of the peroneal nerve were crushed in the sciatic nerve. In the control group, the right sciatic nerve was crushed at the same time that the saphenous, tibial, and sural nerves were transected. Recovery of pain response in the foot was determined by the skin pinch test. Sensory axon elongation rate was measured by the nerve pinch test. The number of myelinated axons was determined in nerve cross sections stained by Azur blue. Recovery of pain sensitivity in the animals of the experimental group was delayed for 2–3 weeks in comparison to the control group. Moreover, the spatial pattern of pain response in the experimental group was irregular, displaying residual regions of insensitive skin which were not present in controls. The elongation rate of regenerating sensory axons in the experimental group was not decreased, and the number of myelinated axons in the peroneal nerves was even about 10% higher than in the control group. Therefore, we assume that the terminal arborization of the neurilemmal tubes pertaining to the former axon sprouts delayed regrowth of sensory axon terminals in the skin.     

Guidance of regenerating motor axons in larval and juvenile bullfrogs

The segmental distribution of regenerating bullfrog motor axons was mapped in advanced tadpoles and juvenile frogs by stimulating selected muscle nerves and recording from the distal ends of the 3 lumbar ventral roots (VRs) that innervate the hindlimb. When motoneurons were axotomized by VR transection, they reestablished their original innervation fields, rarely, if ever, growing beyond the territory normally supplied by their spinal segment. However, when motoneurons were axotomized in the spinal nerves at the level of the hindlimb plexus, some of them regenerated into limb nerves that lay outside the axons' normal segmental boundaries, and many regenerated into the medial femoral cutaneous nerve, a pathway normally limited to sensory axons. These observations suggest that the ultimate destinations of regenerating axons are largely determined by structures the axons encounter as they penetrate the distal nerve stumps. Thus, axons regenerating from a severed VR grow into that root's own distal stump and reinnervate the hindlimb in a manner that is segmentally appropriate; axons transected near the plexus have access to the pathways of sensory, as well as motor, axons in all 3 lumbar segments, and establish innervation fields that are inappropriate for their segment of origin and their motor function.     

Synthesis of myelin proteins and ultrastructural investigations in regenerating rat sciatic nerve

Myelin protein synthesis, as well as ultrastructural and morphometric changes in regenerating peripheral nerve, was studied. Sciatic nerves of rats were crushed unilaterally; sham-operated nerves of the contralateral side served as controls. For the in vivo experiments, rats were killed at selected periods after the nerves were crushed (30, 60, 90, and 120 days); seven days prior to killing, the animals were injected intravenously with L-[4,5-3H]leucine. For the in vitro experiments, proximal and distal segments of sciatic nerve and equivalent sham-operated nerves were labeled with 3H-amino acid mixture 90 days after axotomy. Purified myelin was isolated from nerve segments; specific radioactivity and gel electrophoretic patterns of proteins were analyzed. Cross-sectional electron microscope (EM) preparations of proximal, distal, and contralateral segments of nerves also were examined. Results showed that the incorporation of labeled amino acids into total myelin proteins was enhanced significantly in the distal segment of sciatic nerves at all of the periods of regeneration studied. The yield of myelin protein per mm distal nerve segment increased as regeneration proceeded. The remyelination of fibers early after nerve crush was weak, whereas it gradually attained the normal range 90-120 days after axotomy. Morphometric analysis of myelin sheath thickness of regenerating axons was consistent with the data obtained for myelin protein synthesis.     

The specificity of re-innervation by identified sensory and motor neurons in the leech

Re-innervation of skin and muscle by identified sensory and motor neurons in segmental ganglia of the leech was studied using physiological techniques. After lesions of peripheral nerves, sensory axons which re-innervated the skin always regained sensitivity to their original stimulus modality (touch, pressure or noxious stimuli). Motor neurons invariably re-innervated the appropriate type of body wall muscle, such as longitudinal or circular muscle layers. Both sensory and motor axons usually returned to the appropriate region of the body wall (dorsal, lateral, or ventral) when regenerating after a nerve crush or cut. This capacity was lost, however, when growth along old nerve branches was prevented by evulsing long segments of the nerve. Re-innervation usually occurred by way of growth of new axons all the way the periphery, but in a few cases reconnection with the surviving distal segment of the original axon had taken place. The specificity of reinnervation can be accounted for by a combination of selective growth along appropriate nerve branches and specific interactions with target tissues.     

Schwann cell basal lamina and nerve regeneration

Nerve segments approximately 7 mm long were excised from the predegenerated sciatic nerves of mice, and treated 5 times by repetitive freezing and thawing to kill the Schwann cells. Such treated nerve segments were grafted into the original places so as to be in contact with the proximal stumps. The animals were sacrificed 1, 2, 3, 5, 7 and 10 days after the grafting. The grafts were examined by electron microscopy in the middle part of the graft, i.e. 3-4 mm distal to the proximal end and/or near the proximal and distal ends of the graft. In other instances, the predegenerated nerve segments were minced with a razor blade after repetitive freezing and thawing. Such minced nerves were placed in contact with the proximal stumps of the same nerves. The animals were sacrificed 10 days after the grafting. Within 1-2 days after grafting, the dead Schwann cells had disintegrated into fragments. They were then gradually phagocytosed by macrophages. The basal laminae of Schwann cells, which were not attacked by macrophages, remained as empty tubes (basal lamina scaffolds). In the grafts we examined, no Schwann cells survived the freezing and thawing process. The regenerating axons always grew out through such basal lamina scaffolds, being in contact with the inner surface of the basal lamina (i.e. the side originally facing the Schwann cell plasma membrane). No axons were found outside of the scaffolds. One to two days after grafting, the regenerating axons were not associated with Schwann cells, but after 5-7 days they were accompanied by Schwann cells which were presumed to be migrating along axons from the proximal stumps. Ten days after grafting, proliferating Schwann cells observed in the middle part of the grafts had begun to sort out axons. In the grafts of minced nerves, the fragmented basal laminae of the Schwann cells re-arranged themselves into thicker strands or small aggregations of basal laminae. The regenerating axons, without exception, attached to one side of such modified basal laminae. Collagen fibrils were in contact with the other side, indicating that these modified basal laminae had the same polarity in terms of cell attachment as seen in the ordinary basal laminae of the scaffolds.(ABSTRACT TRUNCATED AT 400 WORDS)     

Axonal elongation through long acellular nerve segments depends on recruitment of phagocytic cells from the near-nerve environment. Electrophysiological and morphological studies in the cat

The distal nerve stump plays a central role in the regeneration of peripheral nerve but the relative importance of cellular and humoral factors is not clear. We have studied this question by freezing the tibial nerve distal to a crush lesion in cat. The importance of constituents from the near-nerve environment was assessed by modification of the contact between the tibial nerve and the environment. Silicone cuffs, containing electrodes for electrophysiological assessment of nerve regeneration, were placed around the tibial nerve distal to the crush site. The interaction between long acellular frozen nerve segments (ANS) and the near-nerve environment was ascertained by breaching the silicone cuff to allow access of cellular or humoral components. Tibial nerves were crushed and frozen for 40 mm and enclosed in nerve cuffs with 0.45-microm holes or 2.0-mm holes to allow access of humoral factors or tissue ingrowth, respectively. In a second set of experiments, tibial nerves were crushed and either frozen for 20+20 mm, leaving a 10 mm segment with viable cells in the center (stepping-stone segment) or frozen for 50 mm. These nerves were enclosed in cuffs with 2.0 mm holes corresponding to the viable nerve segment. The regeneration was monitored electrophysiologically by implanted electrodes and after 2 months the nerves were investigated by light and electron microscopy. The results indicate that soluble substances in the near-nerve environment, such as nutrients, oxygen or tropic substances did not exert any independent beneficial effect on the outgrowing axons. However, phagocytic cells entering the acellular segment from the near-nerve environment were crucial for axonal outgrowth in long ANS.     

Peripheral nerve regeneration is delayed in neuropilin 2-deficient mice

Peripheral nerve transection or crush induces expression of class 3 semaphorins by epineurial and perineurial cells at the injury site and of the neuropilins neuropilin-1 and neuropilin-2 by Schwann and perineurial cells in the nerve segment distal to the injury. Neuropilin-dependent class 3 semaphorin signaling guides axons during neural development, but the significance of this signaling system for regeneration of adult peripheral nerves is not known. To test the hypothesis that neuropilin-2 facilitates peripheral-nerve axonal regeneration, we crushed sciatic nerves of adult neuropilin-2-deficient and littermate control mice. Axonal regeneration through the crush site and into the distal nerve segment, repression by the regenerating axons of Schwann cell p75 neurotrophin receptor expression, remyelination of the regenerating axons, and recovery of normal gait were all significantly slower in the neuropilin-2-deficient mice than in the control mice. Thus, neuropilin-2 facilitates peripheral-nerve axonal regeneration.     

The effect of hyperbaric oxygen treatment on early regeneration of sensory axons after nerve crush in the rat

Abstract The effect of hyperbaric oxygen treatment (HBO) on sensory axon regeneration was examined in the rat. The sciatic nerve was crushed in both legs. In addition, the distal stump of the sural nerve on one side was made acellular and its blood perfusion was compromised by freezing and thawing. Two experimental groups received hyperbaric exposures (2.5 ATA) to either compressed air (pO2 = 0.5 ATA) or 100% oxygen (pO2 = 2.5 ATA) 90 minutes per day for 6 days. Sensory axon regeneration in the sural nerve was thereafter assessed by the nerve pinch test and immunohistochemical reaction to neurofilament. HBO treatment increased the distances reached by the fastest regenerating sensory axons by about 15% in the distal nerve segments with preserved and with compromised blood perfusion. There was no significant difference between the rats treated with different oxygen tensions. The total number of regenerated axons in the distal sural nerve segments after a simple crush injury was not affected, whereas in the nerve segments with compromised blood perfusion treated by the higher pO2, the axon number was about 30% lower than that in the control group. It is concluded that the beneficial effect of HBO on sensory axon regeneration is not dose-dependent between 0.5 and 2.5 ATA pO2. Although the exposure to 2.5 ATA of pO2 moderately enhanced early regeneration of the fastest sensory axons, it decreased the number of regenerating axons in the injured nerves with compromised blood perfusion of the distal nerve stump.     

Collateral sprouting from sensory and motor axons into an end to side attached nerve segment

A nerve segment, sutured end-to-side (ETS) to an intact rat sciatic nerve, becomes invaded by regenerating axons. The origin of these fibres is controversial: it is debated whether or not they represent collateral sprouts from intact axons. Here we demonstrate by double retrograde tracing, using one tracer for the ETS attached segment and another for the sciatic nerve proper double-labelled sensory neurons in 67% of the rats receiving an ETS segments. Double-labelled motor neurons were observed in 11% of the rats. The results show that a nerve segment attached ETS to an intact nerve can induce collateral sprouting of both sensory and motor axons although the extent of such branching may vary with the experimental conditions.     

Preferential reinnervation of motor nerves by regenerating motor axons

Regeneration of axons into inappropriate distal nerve branches may adversely affect functional recovery after peripheral nerve suture. The degree to which motor axons reinnervate sensory nerves, and vice versa, has not been determined. In these experiments, HRP is used to quantify the sensory and motor neurons that reinnervate sensory and motor branches of the rat femoral nerve after proximal severance and repair. Motoneurons preferentially reinnervate the motor branch in juveniles and adults, even if the repair is intentionally misaligned or a gap is imposed between proximal and distal stumps. A specific interaction thus occurs between regenerating motor axons and the Schwann cell tubes that lead to the motor branch. This interaction is independent of mechanical axon alignment.     

Influence of non-neuronal cells on regeneration of the rat sciatic nerve

The ability of the rat sciatic nerve to regenerate into a previously frozen distal nerve segment was studied and compared to regeneration after a crush lesion. The regeneration rate in the frozen segment was 1.9 mm/day, which was approximately half of that observed after a crush lesion (3.3 mm/day). If an unfrozen nerve segment was left intact beyond the frozen section, the rate of regeneration increased to 3.2 mm/day. However, a fresh nerve segment sutured along the frozen segment did not significantly affect the rate of regeneration. Incorporation of [3H]thymidine in the regenerating nerve, analyzed after 1, 3 and 6 days, showed an increased labelling in the frozen segment. This increase spread from the proximal nerve segment into the frozen section. In nerves where a segment was left intact beyond the frozen section, [3H]thymidine incorporation was seen to enter the frozen section from both sides. The spreading of [3H]thymidine incorporation appeared to correlate with the rate of regeneration. However, the same pattern of incorporation could be observed in nerves where regeneration was detained by a transection. The results suggest that Schwann and/or other cells which invade the frozen nerve segment affect the rate of axonal elongation, and that the migration of these cells occurs independently of regenerating fibers.     

4S RNA is transported axonally in normal and regenerating axons of the sciatic nerves of rats

Experiments were designed to determine if following injection of [³H]uridine into the lumbar spinal cord of the rat, [³H]RNA could be demonstrated within axons of the sciatic nerve, and if 4S RNA is the predominant RNA species present in these axons.

In one experiment the left sciatic nerve of a rat was crushed. Two days later 170 μCi of [³H]uridine was injected into the vicinity of the lumbar ventral horn cells. Ten days after injection, rats were sacrificed and sciatic nerves were prepared for autoradiography. Photomicrographs were taken of labeled areas of intact and regenerating nerves and grains were counted over Schwann cells, myelin, axons and other unspecified areas. In both intact and regenerating sciatic nerves more than 20% of the silver grains were associated with motor axons and approximately 40% were found over cytoplasm of Schwann cells surrounding these axons. These data indicate an intra-axonal localization of RNA in sciatic nerve axons, as well as an active transfer of RNA precursors from axons to their surrounding Schwann cells.

In separate studies, the left sciatic nerve was crushed and 10 days later [³H]-uridine was bilaterally injected intraspinally into 6 rats. Four control rats were sacrificed at 14 or 20 days after injection. In the remaining 2 rats the sciatic nerve was cut 14 days after injection and the distal part of the nerve was allowed to degenerate for 6 days before sacrificing the rat. Thus, the distal portion of the nerve contained Schwann cells labeled by axonal transport but lacked intact axons. RNA was isolated from experimental and control nerve segments by hot phenol extraction and ethanol precipitation. RNA species (28S, 18S and 4S) were separated by polyacrylamide gel electrophoresis and radioactivity was measured in a liquid scintillation counter. Control groups had RNA profiles similar to those already described²⁰, with greater than 30% of the radioactivity present as 4S RNA. The proximal portions of nerve taken from the group in which nerves were cut, had a similar amount of radioactivity present as 4S RNA. However, in the distal segments of these nerves (in which the axons had degenerated thus creating an ‘axon-less’ nerve) the amount of radioactivity in the 4S peak decreased to approximately 15% of the total RNA, suggesting that 4S RNA is the predominant if not the only RNA present in these axons. These results strongly indicate that both intact and regenerating sciatic nerves of rats selectively transport 4S RNA along their motor axons.     

The role of laminin, a component of Schwann cell basal lamina, in rat sciatic nerve regeneration within antiserum-treated nerve grafts

Regeneration of the sciatic nerve in transplanted nerve grafts in which laminin was inactivated was examined electron microscopically. Nerve grafts for transplantation were obtained from close cloned donor Wistar rats; 1-cm nerve segments of the sciatic nerve were frozen and thawed to kill the Schwann cells. Control recipient rats received grafts treated with normal rabbit serum to repair the artificially-made complete defect of the right sciatic nerve, and the experimental group of rats received grafts doubly treated with normal serum and rabbit anti-laminin antiserum. In the control grafts regenerating axons grew almost completely through the inside of the basal lamina scaffolds (92%) and adhered to the structure, while in the anti-laminin antiserum treated grafts the axons were present outside (52%) and inside (48%) the scaffolds simultaneously. In this case, the adhesion of axons to the scaffolds was obscure. Axons were associated with and without Schwann cells both inside and outside the basal lamina scaffolds. No unassociated Schwann cells were observed. The maximal number of axons in a 2 mm portion of the antiserum-treated grafts was approximately 250 axons per 100 × 100 μm square and 520 in the control at 15 days. At 30 days, almost the same number of axons was found at the distal (8 mm) portion of both groups. The growth in the former was delayed for 3 days. These results indicate that regenerating peripheral nerve axons may enter the basal lamina scaffolds and grow well because of the neurotrophic function of laminin present at the inner side of Schwann cell basal lamina.     

Acetylcholinesterase and inhibitors: effects upon normal and regenerating nerves of the rat

In peripheral nerves, the function of acetylcholinesterase (AChE) is not related to hydrolysis of acetylcholine. To test for a trophic role, AChE or its inhibitors were administered locally to normal and regenerating nerves of rats. In the normal nerve, neither AChE nor serum albumin affected the cytological pattern of the nerve. BW284c51, a specific inhibitor of AChE, resulted in demyelination, proliferation of Schwann cells and sprouting of axons after 5-7 days. Edrophonium or propidium, other specific inhibitors of AChE, did so to a much lesser extent. Vehicle, and iso-OMPA (inhibitor of pseudocholinesterases) did not affect the cytology of the nerve. Elongation of regenerating axons was evaluated at day 3 post-crush. Native AChE applied distal to the crush reduced the elongation of regenerating axons (- 36%), while serum albumin, heated AChE and filtered AChE did not. BW284c51, edrophonium or propidium enhanced the axonal elongation (33%) when they were administered for 2 days before, but not after, the crush. Iso-OMPA or vehicle administered before or after the crush were not effective. Thus, AChE reduces elongation of regenerating axons, while inhibition of AChE enhances elongation and affects the cytology of the normal nerve as well. We propose that AChE has a trophic role in mammalian peripheral nerves.     

Isolated satellite cells of a peripheral nerve direct the growth of regenerating frog axons

Frog motor axons regenerate and grow back to reinnervate their targets, the original motor end plates, after a lesion. When the cutaneous pectoris muscle is cut away and a segment of peripheral nerve is placed in the vicinity of regenerating axons they turn and grow toward it. This is in marked contrast to the random pattern of axonal outgrowth seen in the absence of a target. The influence on the direction of axonal growth of motor neurons can be produced by a 1-mm segment of nerve satellite cells over a distance of more than 8 mm. The nerve satellite cells have no influence on the direction of growth of the regenerating axons after all the cells in the nerve segment have been killed, leaving only the Schwann cell basal lamina tubes intact. These results show that the cells in the segment of the nerve trunk contain cues that actively direct the growth of motor neurons. Two possible explanations for this effect might be that the cells act indirectly by influencing the organization of the substructure over which axons regenerate or that the nerve satellite cells release a diffusible substance that acts directly on the regenerating axons.     

Induction of myelin genes during peripheral nerve remyelination requires a continuous signal from the ingrowing axon

The effect of a permanent transection on myelin gene expression in a regenerating sciatic nerve and in an adult sciatic nerve was compared to establish the degree of axonal control exerted upon Schwann cells in each population. First, the adult sciatic nerve was crushed, and the distal segment allowed to regenerate. At 12 days post-crush, the sciatic nerve was transected distal to the site of crush to disrupt the Schwann cell-axonal contacts that had reformed. Messenger RNA (mRNA) levels coding for five myelin proteins were assayed in the distal segment of the crush-transected nerve after 9 days and were compared to corresponding levels in the distal segments of sciatic nerves at 21 days post-crush and 21 days post-transection using Northern blot and slot-blot analysis. Levels of mRNAs found in the distal segment of the transected and crush-transected nerve suggested that Schwann cells in the regenerating nerve and in the mature adult nerve are equally responsive to axonal influences. The crush-transected model allowed the genes that were studied to be classified according to their response to Schwann cell-axonal contact. The levels of mRNAs were (1) down-regulated to basal levels (PO and MBP mRNAs), (2) down-regulated to undetectable levels (myelin-associated glycoprotein mRNAs), (3) upregulated (mRNAs encoding 2′3′-cyclic nucleotide phosphodiesterase and β-actin), or (4) not stringently controlled by the removal of Schwann cell-axonal contact (proteolipid protein mRNAs). This novel experimental model has thus provided evidence that the expression of some of the important myelin genes during peripheral nerve regeneration is dependent on continuous signals from the ingrowing axons. © 1993 Wiley-Liss, Inc.     

Diffusion tensor imaging to assess axonal regeneration in peripheral nerves

Development of outcome measures to assess ongoing nerve regeneration in the living animal that can be translated to human can provide extremely useful tools for monitoring the effects of therapeutic interventions to promote nerve regeneration. Diffusion tensor imaging (DTI), a magnetic resonance based technique, provides image contrast for nerve tracts and can be applied serially on the same subject with potential to monitor nerve fiber content. In this study, we examined the use of ex vivo high-resolution DTI for imaging intact and regenerating peripheral nerves in mice and correlated the MRI findings with electrophysiology and histology. DTI was done on sciatic nerves with crush, without crush, and after complete transection in different mouse strains. DTI measures, including fractional anisotropy (FA), parallel diffusivity, and perpendicular diffusivity were acquired and compared in segments of uninjured and crushed/transected nerves and correlated with morphometry. A comparison of axon regeneration after sciatic nerve crush showed a comparable pattern of regeneration in different mice strains. FA values were significantly lower in completely denervated nerve segments compared to uninjured sciatic nerve and this signal was restored toward normal in regenerating nerve segments (crushed nerves). Histology data indicate that the FA values and the parallel diffusivity showed a positive correlation with the total number of regenerating axons. These studies suggest that DTI is a sensitive measure of axon regeneration in mouse models and provide basis for further development of imaging technology for application to living animals and humans.