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Following spinal cord injury (SCI) the degree of functional (motor, autonomous, or sensory) loss correlates with the severity of nervous tissue damage. An imaging technique able to capture non-invasively and simultaneously the complex mechanisms of neuronal loss, vascular damage, and peri-lesional tissue reorganization is currently lacking in experimental SCI studies. Synchrotron X-ray phase-contrast tomography (SXPCT) has emerged as a non-destructive three-dimensional (3D) neuroimaging technique with high contrast and spatial resolution. In this framework, we developed a multi-modal approach combining SXPCT, histology and correlative methods to study neurovascular architecture in normal and spinal level C4-contused mouse spinal cords (C57BL/6J mice, age 2-3 months). The evolution of SCI lesion was imaged at the cell resolution level during the acute (30 min) and subacute (7 day) phases. Spared motor neurons (MNs) were segmented and quantified in different volumes localized at and away from the epicenter. SXPCT was able to capture neuronal loss and blood-brain barrier breakdown following SCI. Three-dimensional quantification based on SXPCT acquisitions showed no additional MN loss between 30 min and 7 days post-SCI. In addition, the analysis of hemorrhagic (at 30 min) and lesion (at 7 days) volumes revealed a high similarity in size, suggesting no extension of tissue degeneration between early and later time-points. Moreover, glial scar borders were unevenly distributed, with rostral edges being the most extended. In conclusion, SXPCT capability to image at high resolution cellular changes in 3D enables the understanding of the relationship between hemorrhagic events and nervous structure damage in SCI.

The topographical neuroanatomy of the human spinal cord (SC) is currently based on the adjacent vertebrae. This morphometric study sought to develop a dataset allowing for statistical analysis of human SC segment characteristics. Overall, 32 human SCs were dissected (18 female and 14 male donors), and individual SC segments were identified. Anterior and posterior lengths, thicknesses and widths were measured by two examiners. Statistical analyses included t-tests, as well as intraclass and Pearson's correlation coefficients. The SC length was significantly shorter in females than males. The cranial (C4) and caudal (T1/T2) limits of the cervical enlargement, along with its maximal width (C6-C7), were identified by combining widths and thicknesses of the segments. The thoracic region, T2 to T12, could be identified using segments widths and thicknesses values. The length of the lumbosacral region, from segments L2 to S5, was particularly stable, independently of SC length and sex. The lumbar enlargement was characterized by a thickness increase between the segments L2 and S1 which reached its maximum at the level of L3, L4, and L5, whereas the width was not significantly increased. From the S2 to S5 segments, width and thickness were equal, with both decreasing of 1 mm per segment. The morphometrical analysis of 32 human SCs provided a dataset allowing for statistical analysis of segmental measures with significant results. A combined approach mostly using widths and thicknesses provided landmarks of potential interest for the localization of SC segments in a clinical MRI setting., (© 2023 American Association of Clinical Anatomists and British Association of Clinical Anatomists.)

Traumatic spinal cord injury (SCI) is a neurologic condition characterized by long-term motor and sensory neurologic deficits as a consequence of an external physical impact damaging the spinal cord. Anatomic MRI is considered the gold-standard diagnostic tool to obtain structural information for the prognosis of acute SCI; however, it lacks functional objective information to assess SCI progression and recovery. In this study, we explored the use of synaptic vesicle glycoprotein 2A (SV2A) PET imaging to detect spinal cord lesions noninvasively after SCI. Methods: Mice ( n = 7) and rats ( n = 8) subjected to unilateral moderate cervical (C5) contusion were euthanized 1 wk after SCI for histologic and autoradiographic ( 3 H-labeled (4 R )-1-[(3-methylpyridin-4-yl)methyl]-4-(3,4,5-trifluorophenyl)pyrrolidin-2-one [UCB-J]) investigation of SV2A levels. Longitudinal 11 C-UCB-J PET/CT imaging was performed in sham ( n = 7) and SCI rats ( n = 8) 1 wk and 6 wk after SCI. Animals also underwent an 18 F-FDG PET scan during the latter time point. Postmortem tissue SV2A analysis to corroborate in vivo PET findings was performed 6 wk after SCI. Results: A significant SV2A loss (ranging from -70.3% to -87.3%; P < 0.0001) was measured at the epicenter of the impact in vitro in both mouse and rat contusion SCI models. Longitudinal 11 C-UCB-J PET imaging detected SV2A loss in SCI rats (-49.0% ± 8.1% at 1 wk and -52.0% ± 12.9% at 6 wk after SCI), with no change observed in sham rats. In contrast, 18 F-FDG PET imaging measured only subtle hypometabolism (-17.6% ± 14.7%). Finally, postmortem 3 H-UCB-J autoradiography correlated with the in vivo SV2A PET findings ( r = 0.92, P < 0.0001). Conclusion: C-UCB-J PET/CT imaging is a noninvasive marker for SV2A loss after SCI. Collectively, these findings indicate that SV2A PET may provide an objective measure of SCI and thus represent a valuable tool to evaluate novel therapeutics. Clinical assessment of SCI with SV2A PET imaging is highly recommended.11 C-UCB-J PET/CT imaging is a noninvasive marker for SV2A loss after SCI. Collectively, these findings indicate that SV2A PET may provide an objective measure of SCI and thus represent a valuable tool to evaluate novel therapeutics. Clinical assessment of SCI with SV2A PET imaging is highly recommended., (© 2022 by the Society of Nuclear Medicine and Molecular Imaging.)

Reactive microgliosis is an important pathological component of neuroinflammation and has been implicated in a wide range of brain diseases including brain tumors, multiple sclerosis, Parkinson's disease, Alzheimer's disease, and schizophrenia. Mapping reactive microglia in-vivo is often performed with PET scanning whose resolution, cost, and availability prevent its widespread use. The advent of diffusion compartment imaging (DCI) to probe tissue microstructure in vivo holds promise to map reactive microglia using MRI scanners. But this potential has never been demonstrated. In this paper, we performed longitudinal DCI in rats that underwent dorsal root axotomy triggering Wallerian degeneration of axons-a pathological process which reliably activates microglia. After the last DCI at 51 days, rats were sacrificed and histology with Iba-1 immunostaining for microglia was performed. The fraction of extra-axonal restricted diffusion from DCI was found to follow the expected temporal dynamics of reactive microgliosis. Furthermore, a strong and significant correlation between this parameter and histological measurement of microglial density was observed. These findings strongly suggest that extra-axonal restricted diffusion is an in-vivo marker of reactive microglia. They pave the way for MRI-based microglial mapping which may be important to characterize the pathogenesis of neurological and psychiatric diseases.

Many closed-form analytical models have been proposed to relate the diffusion-weighted magnetic resonance imaging (DW-MRI) signal to microstructural features of white matter tissues. These models generally make assumptions about the tissue and the diffusion processes which often depart from the biophysical reality, limiting their reliability and interpretability in practice. Monte Carlo simulations of the random walk of water molecules are widely recognized to provide near groundtruth for DW-MRI signals. However, they have mostly been limited to the validation of simpler models rather than used for the estimation of microstructural properties. This work proposes a general framework which leverages Monte Carlo simulations for the estimation of physically interpretable microstructural parameters, both in single and in crossing fascicles of axons. Monte Carlo simulations of DW-MRI signals, or fingerprints, are pre-computed for a large collection of microstructural configurations. At every voxel, the microstructural parameters are estimated by optimizing a sparse combination of these fingerprints. Extensive synthetic experiments showed that our approach achieves accurate and robust estimates in the presence of noise and uncertainties over fixed or input parameters. In an in vivo rat model of spinal cord injury, our approach provided microstructural parameters that showed better correspondence with histology than five closed-form models of the diffusion signal: MMWMD, NODDI, DIAMOND, WMTI and MAPL. On whole-brain in vivo data from the human connectome project (HCP), our method exhibited spatial distributions of apparent axonal radius and axonal density indices in keeping with ex vivo studies. This work paves the way for microstructure fingerprinting with Monte Carlo simulations used directly at the modeling stage and not only as a validation tool., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Recently studies have aimed at developing transcutaneous spinal direct current stimulation (tsDCS) as a non-invasive technique to modulate spinal function in humans. Independent studies evaluating its after-effects on nociceptive or non-nociceptive somatosensory responses have reported comparable effects suggesting that tsDCS impairs axonal conduction of both the spino-thalamic and the medial lemniscus tracts. The present study aimed to better understand how tsDCS affects, in humans, the spinal transmission of nociceptive and non-nociceptive somatosensory inputs. We compared the after-effects of anodal low-thoracic, anodal cervical and sham tsDCS on the perception and brain responses elicited by laser stimuli selectively activating Aδ-thermonociceptors of the spinothalamic system and vibrotactile stimuli selectively activating low-threshold Aβ-mechanoreceptors of the lemniscal system, delivered to the hands and feet. Low-thoracic tsDCS selectively and significantly affected the LEP-N2 wave elicited by nociceptive stimulation of the lower limbs, without affecting the LEP-N2 wave elicited by nociceptive stimulation of the upper limbs, and without affecting the SEP-N2 wave elicited by vibrotactile stimulation of either limb. This selective and segmental effect indicates that the neuromodulatory after-effects of tsDCS cannot be explained by anodal blockade of axonal conduction and, instead, are most probably due to a segmental effect on the synaptic efficacy of the local processing and/or transmission of nociceptive inputs in the dorsal horn., (Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.)

Glioblastoma multiforme (GBM) is the most common primary malignant brain tumor. Despite available therapeutic options, the prognosis for patients with GBM remains very poor. We hypothesized that the intra-operative injection of a photopolymerizable hydrogel into the tumor resection cavity could sustain the release of the anti-cancer drug paclitaxel (PTX) encapsulated in poly (lactic-co-glycolic acid) (PLGA) nanoparticles and prevent GBM recurrence. The tumor was resected 13 days after implantation and a pre-gel solution composed of polyethylene glycol dimethacrylate (PEG-DMA) polymer, a photoinitiator and PTX-loaded PLGA nanoparticles (PTX PLGA-NPs) was injected into the tumor resection cavity. A solid gel filling the whole cavity was formed immediately by photopolymerization using a 400 nm light. PTX in vitro release study showed a burst release (11%) in the first 8 h and a sustained release of 29% over a week. In vitro, U87 MG cells were sensitive to PTX PLGA-NPs with IC 50 level of approximately 0.010 μg/mL. The hydrogel was well-tolerated when implanted in the brain of healthy mice for 2 and 4 months. Administration of PTX PLGA-NPs-loaded hydrogel into the resection cavity of GBM orthotopic model lead to more than 50% long-term survival mice (150 days) compared to the control groups (mean survival time 52 days). This significant delay of recurrence is very promising for the post-resection treatment of GBM., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Although brain tumours have been documented and recorded since the nineteenth century, 2016 marked 90 years since Percival Bailey and Harvey Cushing coined the term "glioblastoma multiforme". Since that time, although extensive developments in diagnosis and treatment have been made, relatively little improvement on prognosis has been achieved. The resilience of GBM thus makes treating this tumour one of the biggest challenges currently faced by neuro-oncology. Aggressive and robust development, coupled with difficulties of complete resection, drug delivery and therapeutic resistance to treatment are some of the main issues that this nemesis presents today. Current treatments are far from satisfactory with poor prognosis, and focus on palliative management rather than curative intervention. However, therapeutic research leading to developments in novel treatment stratagems show promise in combating this disease. Here we present a review on GBM, looking at the history and advances which have shaped neurosurgery over the last century that cumulate to the present day management of GBM, while also exploring future perspectives in treatment options that could lead to new treatments on the road to a cure.

We hypothesized that local delivery of GDNF in spinal cord lesion via an injectable alginate hydrogel gelifying in situ would support spinal cord plasticity and functional recovery. The GDNF release from the hydrogel was slowed by GDNF encapsulation in microspheres compared to non-formulated GDNF (free GDNF). When injected in a rat spinal cord hemisection model, more neurofilaments were observed in the lesion when the rats were treated with free GDNF-loaded hydrogels. More growing neurites were detected in the tissues surrounding the lesion when the animals were treated with GDNF microsphere-loaded hydrogels. Intense GFAP (astrocytes), low βIII tubulin (neural cells) and RECA-1 (endothelial cells) stainings were observed for non-treated lesions while GDNF-treated spinal cords presented less GFAP staining and more endothelial and nerve fiber infiltration in the lesion site. The animals treated with free GDNF-loaded hydrogel presented superior functional recovery compared with the animals treated with the GDNF microsphere-loaded hydrogels and non-treated animals., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Brain responses to the activation of C-fibres are obtained only if the co-activation of Aδ-fibres is avoided. Methods to activate C-fibres selectively have been proposed, but are unreliable or difficult to implement. Here, we propose an approach combining a new laser stimulator to generate constant-temperature heat pulses with an adaptive paradigm to maintain stimulus temperature above the threshold of C-fibres but below that of Aδ-fibres, and examine whether this approach can be used to record reliable C-fibre laser-evoked brain potentials. Brief CO2 laser stimuli were delivered to the hand and foot dorsum of 10 healthy subjects. The stimuli were generated using a closed-loop control of laser power by an online monitoring of target skin temperature. The adaptive algorithm, using reaction times to distinguish between late detections indicating selective activation of unmyelinated C-fibres and early detections indicating co-activation of myelinated Aδ-fibres, allowed increasing the likelihood of selectively activating C-fibres. Reliable individual-level electroencephalogram (EEG) responses were identified, both in the time domain (hand: N2: 704 ± 179 ms, P2: 984 ± 149 ms; foot: N2: 1314 ± 171 ms, P2: 1716 ± 171 ms) and the time-frequency (TF) domain. Using a control dataset in which no stimuli were delivered, a Receiver Operating Characteristics analysis showed that the magnitude of the phase-locked EEG response corresponding to the N2-P2, objectively quantified in the TF domain, discriminated between absence vs presence of C-fibre responses with a high sensitivity (hand: 85%, foot: 80%) and specificity (hand: 90%, foot: 75%). This approach could thus be particularly useful for the diagnostic workup of small-fibre neuropathies and neuropathic pain., (Copyright © 2013. Published by Elsevier B.V.)

Objectives: To retrospectively identify morphological and physiological post-operative magnetic resonance imaging (MRI) characteristics predictive of glioblastoma recurrences after gross total resection (gross-TR)., Methods: Resection margins of 24 glioblastoma were analysed immediately post-operatively (MRI ≤ 2 h) and early post-operatively (24 h ≤ MRI ≤ 48 h), and subdivided into areas with and without subtle contrast enhancement previously considered non-specific. On follow-up MRI, tumour regrowth areas were subdivided according to recurrence extent (focally/extended) and delay (≤6 and ≥12 months). Co-registration of pre-operative, immediately post-operative and early post-operative MRI with the first follow-up MRI demonstrating recurrence authorised their morphological (contrast enhancements) and physiological (rCBV) characterisation., Results: Morphologically, on immediately post-operative MRI, micro-nodular and frayed enhancements correlate significantly with early recurrences (≤6 months). After gross-TR the absence of these enhancements is associated with a significant increase in progression-free survival (61 vs 15 weeks respectively) and overall survival (125 vs 51 weeks respectively). Physiologically, areas with a future focal recurrence have a trend toward higher rCBV than other areas., Conclusion: Immediately post-operative topography of micro-nodular and frayed enhancements is suggestive of recurrence location and delay. Absence of such enhancements is associated with a fourfold increase in progression-free survival and a 2.5-fold increase in overall survival., Key Points: • Immediately post-operative MRI reveals contrast enhancement after glioblastoma gross total resection. • Immediately post-operative micro-nodular and frayed enhancement correlate with early recurrence. • Absence of micro-nodular/frayed enhancement is associated with 61 weeks' progression-free survival. • Absence of micro-nodular/frayed enhancement is associated with 125 weeks' overall survival.

We assessed feasibility, safety, and timing of an original intraoperative MR procedure in 3 cases of resection of spinal cord glioma by using a clinical 3T MR system connected to an adjacent operating room in a design being coined "twin" or "dual" MR-operating room suite.

Objective: A twin neurosurgical magnetic resonance imaging (MRI) suite with 3-T intraoperative MRI (iMRI) was developed to be available to neurosurgeons for iMRI and for independent use by radiologists., Methods: The suite was designed with one area dedicated to neurosurgery and the other to performing MRI under surgical conditions (sterility and anesthesia). The operating table is motorized, enabling transfer of the patient into the MRI system. These two areas can function independently, allowing the MRI area to be used for nonsurgical cases. We report the findings from the first 21 patients to undergo scheduled neurosurgery with iMRI in this suite (average age, 51 +/- 24 yr; intracranial tumor, 18 patients; epilepsy surgery, 3 patients)., Results: Twenty-six iMRI examinations were performed, 3 immediately before surgical incision, 9 during surgery (operative field partially closed), and 14 immediately postsurgery (operative field fully closed but patient still anesthetized and draped). Minor technical dysfunctions prolonged 10 iMRI procedures; however, no serious iMRI-related incidents occurred. Twenty-three iMRI examinations took an average of 78 +/- 20 minutes to perform. In three patients, iMRI led to further tumor resection because removable residual tumor was identified. Complete tumor resection was achieved in 15 of the 18 cases., Conclusion: The layout of the new complex allows open access to the 3-T iMRI system except when it is in use under surgical conditions. Three patients benefited from the iMRI examination to achieve total resection. No permanent complications were observed. Therefore, the 3-T iMRI is feasible and appears to be a safe tool for intraoperative surgical planning and assessment.

Most studies on the origin of oligodendrocyte lineage have been performed in the spinal cord. By contrast, molecular mechanisms that regulate the appearance of the oligodendroglial lineage in the brain have not yet attracted much attention. We provide evidence for three distinct sources of oligodendrocytes in the mouse telencephalon. In addition to two subpallial ventricular foci, the anterior entopeduncular area and the medial ganglionic eminence, the rostral telencephalon also gives rise to oligodendrocytes. We show that oligodendrocytes in the olfactory bulb are generated within the rostral pallium from ventricular progenitors characterized by the expression of PLP: We provide evidence that these Plp oligodendrocyte progenitors do not depend on signal transduction mediated by platelet-derived growth factor receptors (PDGFRs), and therefore propose that they belong to a different lineage than the PDGFRalpha-expressing progenitors. Moreover, induction of oligodendrocytes in the telencephalon is dependent on sonic hedgehog signaling, as in the spinal cord. In all these telencephalic ventricular territories, oligodendrocyte progenitors were detected at about the same developmental stage as in the spinal cord. However, both in vivo and in vitro, the differentiation into O4-positive pre-oligodendrocytes was postponed by 4-5 days in the telencephalon in comparison with the spinal cord. This delay between determination and differentiation appears to be intrinsic to telencephalic oligodendrocytes, as it was not shortened by diffusible or cell-cell contact factors present in the spinal cord.

In vertebrates, interneurons of the olfactory bulb (OB) are generated postnatally and throughout life at the subventricular zone of the forebrain. The neuronal precursors migrate tangentially through the forebrain using a well defined pathway, the rostral migratory stream (RMS), and a particular mode of migration in a chain-like organization. A severe size reduction of the OB represents the most striking morphological phenotype in neural cell adhesion molecule (NCAM)-deficient mice. This defect has been traced back to a migration deficit of the precursors in the RMS and linked to the lack of the polysialylated form of NCAM. In this study we investigate the morphological alterations and functional properties of the RMS in mice totally devoid of all isoforms of NCAM and polysialic acid (PSA). We show that a morphologically altered, but defined and continuous pathway exists in mutants, and we present in vivo and in vitro evidence that PSA-NCAM in the RMS is not essential for the formation and migration of chains. Instead, we find a massive gliosis associated with the formation of membrane specializations in a heterotypic manner, linking precursors to astrocytes. This finding and the over-representation and defasciculation of axons in the pathway suggest that important interactions between migrating cells and their stationary environment are perturbed in the mutants. Finally, we used transplantation experiments to demonstrate that lack of PSA-NCAM leads to a decrease but not a total blockade of migration and demonstrate that the mutant RMS is functional in transporting normal neuronal precursors to the OB.

The subventricular zone of the adult mammalian forebrain contains progenitor cells that, by migrating along a restricted pathway called the 'rostral migratory stream' (RMS), add new neurons to the olfactory bulb throughout life. To determine the influence of the olfactory bulb on the development of these progenitor cells, we performed lesions that interrupt this pathway and separate the olfactory bulb from the rest of the forebrain. By labelling cells born at several survival times after the lesions with the thymidine analogue bromodeoxyuridine (BrdU), we found that disconnection from the bulb influences the rate of BrdU incorporation by the progenitor cells. The number of labelled cells in lesioned mice was almost half that found in control mice. In the disconnected migratory pathway, the number of neurons expressing calretinin was increased indicating that neuronal differentiation was enhanced: newly born neurons occurred within and around the RMS, most of them expressed calretinin and left the pathway starting about 2 weeks after the lesion. Thereafter, these neurons preserving their phenotype, spread for long distances, and accumulated ectopically in dorsal regions of the anterior olfactory nucleus and the frontal cortex. Finally, transplantation of adult subventricular cells into the lesioned pathway showed that the lesion neither prevents neuronal migration nor alters its direction. Thus, although the olfactory bulb appears to regulate the pace of the developmental processes, its disconnection does not prevent the proliferation, migration and phenotypic acquisition of newly generated bulbar interneurons that, since they cannot reach their terminal domains, populate some precise regions of the lesioned adult forebrain.

Neural progenitors are thought to be multipotent cells whose adult phenotype is determined by extrinsic influences acting during and immediately after their last mitosis. To test this hypothesis, postnatal cerebellar precursor cells were placed in the heterochronic cellular environment of the embryonic mouse cerebellar anlage and the resulting phenotypes were determined. To identify the cells arising from postnatal precursors, tissue fragments taken from 3- to 8-day-old cerebellum of several transgenic mouse lines (each expressing the lacZ reporter gene in different sets of neuronal populations) were mixed with fragments taken from the wild-type cerebellar primordium of 12- or 13-day-old embryos. The fragments were dissociated and grafted into the cerebellum of adult mice. The phenotype acquired by postnatal precursors in the mixed grafts was determined by their morphology and ultrastructural features and by the expression of specific markers. Only two adult phenotypes were generated by these precursors: granule cells and molecular layer interneurons. Most granule cells were well integrated in the trilaminated cortex of the graft, being positioned in their proper layer both during development and after complete maturation. By contrast, basket and stellate cells were always ectopic, remaining outside the molecular layer. These results indicate that at least two distinct progenitor cells are present in the postnatal cerebellar cortex under the experimental conditions of this study. Both progenitors appear to be strictly specified at the time of grafting, and neither their identify nor the expression of their major distinctive features are significantly influenced by local signals emerging from the cellular environment of the embryonic cerebellar anlage.

To gain insight into cellular and molecular mechanisms subserving neuronal cell migration in the adult mouse forebrain, we have first investigated the cellular composition of the subventricular zone-olfactory bulb pathway (SVZ-OB). The pathway was essentially composed of cells with neuronal and astrocytic identities, neuronal cells being four times more numerous than astrocytes. Neuronal cells (precursors and some young postmitotic neurons) formed continuous cellular strands of migratory cells from the anterior horn of the lateral ventricle to the olfactory bulb. These chains of migrating cells moved within channels formed by the processes of a special subpopulation of astrocytes. The neuronal cells expressed the embryonic form of polysialic acid neural cell adhesion molecule, and the astrocytes were tenascin-C positive, thus preserving an embryonic cellular environment. Through transplantation experiments, the second part of this study attempted to analyze the functional properties of the adult SVZ-OB pathway. Early postnatal (P2-13) cerebellar progenitor cells, taken from a transgenic mouse line in which cerebellar granule cells and molecular layer interneurons (basket/stellate cells) expressed the reporter gene lacZ, were implanted in the SVZ-OB pathway of adult wild-type mice. Unlike grafted SVZ cells that migrate all along the pathway, none of the cerebellar precursors reached the olfactory bulb, although some of them were able to migrate along the caudal one-third of the pathway. The majority (over 67%) of the migrating cells were progenitors that acquired the phenotype of basket/stellate cells. Granule cell progenitors and most granule cells did not survive transplantation. These results show that the adult SVZ-OB pathway is not a "passive generic guidance" for all classes of premigratory neurons. From the two types of grafted cerebellar progenitors, only those with migratory capability and that do not follow radial glial axes are able to translocate along the SVZ-OB pathway. Furthermore, the basket/stellate cell progenitors are specified at the time of grafting: Neither their identity nor the pace of expression of their major distinctive features are influenced by local signals emanating from the adult forebrain.

Regeneration of severed central axons is supposed to depend on two factors: a permissive local environment and the particular intrinsic properties of axotomized neurones. To assess the role of each of these factors in axonal regeneration, the capability of two particular axon populations of the adult mouse cerebellum to grow into target-specific (cerebellum) and target-unspecific (neocortex) embryonic grafts was determined. Purkinje cell and inferior olivary axons were transected by passing a microscalpel through the axial white matter of the cerebellar folia, particularly those of the anterior lobe. Immediately after the injury, solid transplants were placed in the lesion cavity. Purkinje cell axons were labelled by using anticalbindin immunocytochemistry, and olivocerebellar fibres were visualized by biotinylated dextran amine anterograde axonal tracing. Following axotomy, Purkinje cell axons appeared as thickened processes ending with large terminal clubs. Their morphology and number did not change up to the longest survival time considered (2 months), thereby confirming previous demonstrations that Purkinje cells survive axon injury (I. Dusart and C. Sotelo, 1994, J. Comp. Neurol. 347:211-232). Inferior olivary axons were thinner and bore smaller terminal bulbs. When embryonic cerebellar grafts, containing cortical and deep nuclear precursors, were placed close to the injured axons, olivocerebellar fibres vigorously regenerated into the transplants and ended in new climbing fibres along the dendrites of grafted Purkinje cells. By contrast, host Purkinje cell axons never showed any outgrowth towards the graft. Similarly, these axons failed to regenerate into grafts containing solely the rostromedial portion of the cerebellar anlage, mostly consisting of deep nuclear neurones, their main targets. Comparable results were obtained by transplanting embryonic neocortical tissue: inferior olivary axons also regenerated into the grafts, although with distinct terminal arbours without the climbing fibre phenotype, whereas Purkinje cell axons always failed to grow. These results provide the first direct demonstration that severed inferior olivary axons are able to regenerate. In addition, they show that the growth-permissive/-promoting conditions created by embryonic nervous tissue are not sufficient to induce the regeneration of every axonal type and allow us to hypothesise that successful regeneration depends on the interplay between environmental cues and intrinsic properties of the axotomized neurones.

The effects of target loss on adult axonal arbors were investigated by comparing the morphological changes of adult climbing fibers in several mutant mouse strains where Purkinje cells slowly degenerate (namely, Lurcher, nervous, Purkinje cell degeneration, and tambaleante), with those occurring after a fast Purkinje cell death induced by mechanical lesions of the adult mouse cerebellum. In each of the different mutations, Purkinje cells displayed distinctive structural modifications. However, a set of regressive changes common to all strains could be disclosed, mostly dendritic atrophy and a progressive axonal retraction with the hypertrophy of recurrent collaterals. Climbing fibers that contacted such degenerating neurons also showed abnormal morphological features, consisting in the presence of extensive perisomatic plexuses, whereas peridendritic branches were atrophic or absent. In Lurcher mice, target-deprived climbing fibers were strictly confined around the granular-molecular layer interface and never penetrated into the molecular layer. Similar terminal plexuses at the level of the former Purkinje cell layer were observed in the other mutants. However, in the latter cases, atrophic terminal arbors were also present in the molecular layer, being confined to the deep portions in nervous, while spanning its whole extent in Purkinje cell degeneration and tambaleante mice. Following mechanical lesions, atrophic target-deprived climbing fibers were exclusively located in the molecular layer. In addition, some of the Purkinje cells that survived after the injury displayed regressive modifications similar to those observed in mutant mice, and their climbing fibers were characterized by perisomatic plexuses. These results show that the normal relationship between the climbing fiber and its Purkinje cell is already disrupted during the slow degeneration of the target neuron. As a consequence, the phenotypic pattern of target-deprived climbing fibers reflects the preceding interactions with their postsynaptic neurons and it is determined by the onset time and progression rate of Purkinje cell degeneration.