Human pluripotent stem cell-derived inner ear organoids recapitulate otic development in vitro.

Our molecular understanding of the early stages of human inner ear development has been limited by the difficulty in accessing fetal samples at early gestational stages. As an alternative, previous studies have shown that inner ear morphogenesis can be partially recapitulated using induced pluripotent stem cells directed to differentiate into inner ear organoids (IEOs). Once validated and benchmarked, these systems could represent unique tools to complement and refine our understanding of human otic differentiation and model developmental defects. Here, we provide the first direct comparisons of the early human embryonic otocyst and fetal sensory organs with human IEOs. We use multiplexed immunostaining and single-cell RNA-sequencing to characterize IEOs at three key developmental steps, providing a new and unique signature of in vitro-derived otic placode, epithelium, neuroblasts and sensory epithelia. In parallel, we evaluate the expression and localization of crucial markers at these equivalent stages in human embryos. Together, our data indicate that the current state-of-the-art protocol enables the specification of bona fide otic tissue, supporting the further application of IEOs to inform inner ear biology and disease.

The effects of creatine supplementation on striatal neural progenitor cells depend on developmental stage.

Transplantation of neural progenitor cells (NPCs) is a promising experimental therapy for Huntington's disease (HD). The variables responsible for the success of this approach, including selection of the optimal developmental stage of the grafted cells, are however largely unknown. Supporting cellular energy metabolism by creatine (Cr) supplementation is a clinically translatable method for improving cell transplantation strategies. The present study aims at investigating differences between early (E14) and late (E18) developmental stages of rat striatal NPCs in vitro. NPCs were isolated from E14 and E18 embryos and cultured for 7 days with or without Cr [5 mM]. Chronic treatment significantly increased the percentage of GABA-immunoreactive neurons as compared to untreated controls, both in the E14 (170.4 ± 4.7 %) and the E18 groups (129.3 ± 9.3 %). This effect was greater in E14 cultures (p < 0.05). Similarly, short-term treatment for 24 h resulted in increased induction (p < 0.05) of the GABA-ergic phenotype in E14 (163.0 ± 10.4 %), compared to E18 cultures (133.3 ± 9.5 %). Total neuronal cell numbers and general viability were not affected by Cr (p > 0.05). Protective effects of Cr against a metabolic insult were equal in E14 and E18 NPCs (p > 0.05). Cr exposure promoted morphological differentiation of GABA-ergic neurons, including neurite length in both groups (p < 0.05), but the number of branching points was increased only in the E18 group (p < 0.05). Our results demonstrate that the role of Cr as a GABA-ergic differentiation factor depends on the developmental stage of striatal NPCs, while Cr-mediated neuroprotection is not significantly influenced. These findings have potential implications for optimizing future cell replacement strategies in HD.

Enhanced proliferation and dopaminergic differentiation of ventral mesencephalic precursor cells by synergistic effect of FGF2 and reduced oxygen tension.

Effective numerical expansion of dopaminergic precursors might overcome the limited availability of transplantable cells in replacement strategies for Parkinson's disease. Here we investigated the effect of fibroblast growth factor-2 (FGF2) and FGF8 on expansion and dopaminergic differentiation of rat embryonic ventral mesencephalic neuroblasts cultured at high (20%) and low (3%) oxygen tension. More cells incorporated bromodeoxyuridine in cultures expanded at low as compared to high oxygen tension, and after 6 days of differentiation there were significantly more neuronal cells in low than in high oxygen cultures. Low oxygen during FGF2-mediated expansion resulted also in a significant increase in tyrosine hydroxylase-immunoreactive (TH-ir) dopaminergic neurons as compared to high oxygen tension, but no corresponding effect was observed for dopamine release into the culture medium. However, switching FGF2-expanded cultures from low to high oxygen tension during the last two days of differentiation significantly enhanced dopamine release and intracellular dopamine levels as compared to all other treatment groups. In addition, the short-term exposure to high oxygen enhanced in situ assessed TH enzyme activity, which may explain the elevated dopamine levels. Our findings demonstrate that modulation of oxygen tension is a recognizable factor for in vitro expansion and dopaminergic differentiation of rat embryonic midbrain precursor cells.

Functional effect of FGF2- and FGF8-expanded ventral mesencephalic precursor cells in a rat model of Parkinson's disease.

Ventral mesencephalic (VM) precursor cells are of interest in the search for transplantable dopaminergic neurons for cell therapy in Parkinson's disease (PD). In the present study we investigated the survival and functional capacity of in vitro expanded, primary VM precursor cells after intrastriatal grafting to a rat model of PD. Embryonic day 12 rat VM tissue was mechanically dissociated and cultured for 4 or 8 days in vitro (DIV) in the presence of FGF2 (20 ng/ml), FGF8 (20 ng/ml) or without mitogens (control). Cells were thereafter differentiated for 6 DIV by mitogen withdrawal and addition of serum. After differentiation, significantly more tyrosine hydroxylase-immunoreactive (TH-ir), dopamine-producing neurons were found in FGF2- and FGF8-expanded cultures compared to controls. Moreover, expansion for 4 DIV resulted in significantly more TH-ir cells than expansion for 8 DIV both for FGF2 (2.4 fold; P<0.001) and FGF8 (3.8 fold; P<0.001) treated cultures. The functional potential of the expanded cells (4 DIV) was examined after grafting into striatum of aged 6-hydroxydopamine-lesioned rats. Amphetamine-induced rotations performed 3, 6 and 9 weeks postgrafting revealed that grafts of FGF2-expanded cells induced a significantly faster and better functional recovery than grafts of FGF8-expanded cells or control cells (P<0.05 for both). Grafts of FGF2-expanded cells also contained significantly more TH-ir cells than grafts of FGF8-expanded cells (P<0.05) or control cells (P<0.01). In conclusion, FGF2-mediated pregrafting expansion of primary VM precursor cells considerably improves dopaminergic cell survival and functional restoration in a rat model of PD.

Psychosocial and neurocognitive performance after spontaneous nonaneurysmal subarachnoid hemorrhage related to the APOE-epsilon4 genotype: a prospective 5-year follow-up study.

OBJECT: In this study, the authors prospectively evaluated long-term psychosocial and neurocognitive performance in patients suffering from nonaneurysmal, nontraumatic subarachnoid hemorrhage (SAH) and investigated the association between the APOE-epsilon4 genotype and outcome in these patients. METHODS: All patients admitted to the authors' institution between January 2001 and January 2003 with spontaneous nonaneurysmal SAH were prospectively examined (mean follow-up 59.8 months). The APOE genotype was determined in all patients by polymerase chain reaction from a blood sample. Of the 30 patients included in this study, 11 were carriers of the epsilon4 allele. RESULTS: All patients showed a good recovery and regained full independence with no persisting neurological deficits. The patients with the epsilon4 allele, however, scored significantly higher on the Beck Depression Inventory (22.1 +/- 6.3 vs 14.1 +/- 5.1). At follow-up, depression was more persistent in the group with the epsilon4 allele compared with the group that lacked the allele. This finding reached statistical significance (p < 0.05). Selective attention was impaired in all patients during the first year of follow-up, with an earlier recovery noted in the patients without the epsilon4 allele. Moreover, there was a tendency toward a linear relationship between the Beck Depression Inventory and the d2 Test of Attention. Two patients who carried the epsilon4 allele did not return to their employment even after 5 years. CONCLUSIONS: The findings in this study suggest that the APOE genotypes may be associated with the psychosocial and neurocognitive performance after spontaneous nonaneurysmal SAH, even in the absence of neurological impairment. Physicians should consider patient genotype in assessing the long-term consequences of nonaneurysmal SAH.

Restorative neuroscience: concepts and perspectives.

There is increasing interest in the search for therapeutic options for diseases and injuries of the central nervous system (CNS), for which currently no effective treatment strategies are available. Replacement of damaged cells and restoration of function can be accomplished by transplantation of cells derived from different sources, such as human foetal tissue, genetically modified cell lines, embryonic or somatic stem cells. Preclinical and clinical trials have shown promising results in neurodegenerative disorders, like Parkinson's and Huntington's disease, but also ischaemic stroke, intracerebral haemorrhage, demyelinating disorders, epilepsy and traumatic lesions of the brain and spinal cord. Other studies have focused on finding new ways to activate and direct endogenous repair mechanisms in the CNS, eg, by exposure to specific neuronal growth factors or by inactivating inhibitory molecules. Neuroprotective drugs may offer an additional tool for improving neuronal survival in acute or chronic CNS diseases. Importantly however, a number of scientific issues need to be addressed in order to permit the introduction of these experimental techniques in the wider clinical setting.

Cell replacement therapy for intracerebral hemorrhage.

Intracerebral hemorrhage (ICH), for which no effective treatment strategy is currently available, constitutes one of the most devastating forms of stroke. As a result, developing therapeutic options for ICH is of great interest to the medical community. The 3 potential therapies that have the most promise are cell replacement therapy, enhancing endogenous repair mechanisms, and utilizing various neuroprotective drugs. Replacement of damaged cells and restoration of function can be accomplished by transplantation of cells derived from different sources, such as embryonic or somatic stem cells, umbilical cord blood, and genetically modified cell lines. Early experimental data showing the benefits of cell transplantation on functional recovery after ICH have been promising. Nevertheless, several studies have focused on another therapeutic avenue, investigating novel ways to activate and direct endogenous repair mechanisms in the central nervous system, through exposure to specific neuronal growth factors or by inactivating inhibitory molecules. Lastly, neuroprotective drugs may offer an additional tool for improving neuronal survival in the perihematomal area. However, a number of scientific issues must be addressed before these experimental techniques can be translated into clinical therapy. In this review, the authors outline the recent advances in the basic science of treatment strategies for ICH.

Combination of cell transplantation and glial cell line-derived neurotrophic factor-secreting encapsulated cells in Parkinson's disease.

A major limitation of cell transplantation for Parkinson's disease (PD) is the mediocre survival of the grafted cells. Facilitating graft survival may improve the functional outcomes of the transplanted cells. Here, we discuss our observations that combination of rat fetal ventral mesencephalic (VM) tissue and encapsulated cells that secrete glial cell line-derived neurotrophic factor (GDNF) enhanced graft function in an animal model of PD. We described significant 2-fold increase in the number of tyrosine hydroxylase immunoreactive (TH-ir) cells per graft, as well as 1.7-fold and 9-fold increments in TH-ir fiber outgrowth into the host brain and toward the capsule with combined transplants and GDNF capsules as opposed to the VM transplants and mock-capsule group. These findings demonstrate that encapsulated GDNF-secreting cells improve graft survival that may optimize functional benefits for the treatment of PD.

Neurotrophic factor-based strategies to enhance survival and differentiation of neural progenitor cells toward the dopaminergic phenotype.

Parkinson's disease (PD) is a neurodegenerative disorder that presents with hallmark clinical symptoms of tremor at rest, bradykinesia, and muscle rigidity. Stem cell therapy has emerged as an experimental treatment for PD. However, optimizing the cell culture condition that allows enhanced survival and differentiation of cells toward the dopaminergic phenotype remains a logistical challenge. Here, we discuss the utility of a combination of neurotrophin-4/5 (NT-4/5) and glial cell line-derived neurotrophic factor (GDNF) in increasing the dopaminergic phenotypic expression of rat ventral mesencephalic (VM) tissue. Using organotypic explant cultures of fetal human ventral mesencephalon, we observed that NT-4/5 and GDNF as single factors, or in combination on DAergic neurons, increased survival and number of tyrosine hydroxylase immunoreactive neurons as well as the dopamine content in the culture medium. The application of specific neurotrophic factors, such as NT-4/5 and GDNF, as cell culture supplements or as adjunctive therapy to cell transplantation may achieve improved functional outcomes when contemplating cell-based regenerative medicine for PD.

A Combination of NT-4/5 and GDNF Is Favorable for Cultured Human Nigral Neural Progenitor Cells.

Idiopathic Parkinson's disease (PD) is a progressive neurodegenerative disorder, clinically manifested by cardinal motor symptoms including tremor at rest, bradykinesia, and muscle rigidity. Transplantation of dopaminergic (DAergic) neurons is an experimental therapy for PD, however, it is limited by suboptimal integration and low survival of grafts. Pretreatment of donor tissue may offer a strategy to improve properties of transplanted DAergic neurons and thereby clinical outcome. We have previously shown that a combination of neurotrophin-4/5 (NT-4/5) and glial cell line-derived neurotrophic factor (GDNF) demonstrated additive effects on rat ventral mesencephalic (VM) tissue. The present study investigated the effects of NT-4/5 and GDNF as single factors, or in combination on DAergic neurons, in organotypic explant cultures of fetal human ventral mesencephalon. For that purpose, free-floating roller-tube cultures were prepared from VM and the equally sized pieces grown for 1 week in the presence or absence of neurotrophic factors. Both neurotrophic factors increased dopamine content in the culture medium and in the number of tyrosine hydroxylase immunoreactive neurons, most prominently after combined GDNF + NT-4/5 treatment. Culture volumes did not differ between groups while content of lactate dehydrogenase in the culture medium was moderately reduced in all treated groups. In conclusion, we identified that a combination of GDNF and NT-4/5 robustly promoted differentiation and survival of human fetal VM DAergic neurons, an observation with potential promising impact for cell replacement approaches in PD.

Experimental intracerebral hematoma in the rat.

PURPOSE: Current models of intracerebral hematoma are difficult to use for neurotransplantation studies because of high mortality and important variations of morphology, size and location of blood deposits. We propose a modification of the autologous blood infusion technique in rats to reduce these limitations. METHODS: The modification consisted in a mechanical microlesion preceding blood infusion. A canula was stereotactically introduced into the striatum of adult rats. Subsequently, a parenchyma lesion was created by a rotating microcatheter coaxially inserted through the canula, followed by slow infusion of 30 mul autologous blood during 5 minutes. Controls included canula insertion only and canula + microlesion. Hematoma volume/morphology were quantified and the animals behaviorally analysed using standardized tests. RESULTS: Surgical mortality was 0/54 rats. One animal died during follow-up. Hematoma volume was constant and significantly higher (15.20 +/- 0.60 mm;3) than control lesions (canula: 0.11 +/- 0.01 mm;3; canula + trauma: 0.51 +/- 0.01 mm;3). Hematoma edges were sharply delineated and the perihematomal region histologically preserved. Rats with hematoma showed initially a reduced spontaneous rotational behaviour. They also showed persisting deficits of forelimb placing ability. CONCLUSIONS: The advantages of this model include a systematic control of all steps of hematoma production, high reproducibility of volume, size, and location of blood deposits, preservation of perihematomal brain tissue, and quantifiable neurological deficits.

Antagonization of the Nogo-Receptor 1 Enhances Dopaminergic Fiber Outgrowth of Transplants in a Rat Model of Parkinson's Disease.

Intrastriatal transplantation of fetal human ventral mesencephalic dopaminergic neurons is an experimental therapy for patients suffering from Parkinson's disease. The success of this approach depends on several host brain parameters including neurotrophic factors and growth inhibitors that guide survival and integration of transplanted neurons. While the potential of neurotrophic factors has been extensively investigated, repression of growth inhibitors has been neglected, despite the significant effects reported in various CNS injury models. Recently, we demonstrated that infusion of neutralizing antibodies against Nogo-A into the lateral ventricles of hemi-parkinsonian rats significantly enhanced graft function. Since the Nogo-receptor 1 also interacts with other neurite growth inhibitors, we investigated whether a direct antagonization of the receptor would result in more robust effects. Therefore, rats with unilateral striatal 6-hydroxydopamine lesions were grafted with ventral mesencephalic tissue in combination with intraventricular infusions of the Nogo-receptor 1 antagonist NEP1-40. Transplanted rats receiving saline infusions served as controls. To test whether NEP1-40 treatment alone affects the remaining dopaminergic striatal fibers, rats with unilateral striatal 6-hydroxydopamine lesions were infused with NEP1-40 or saline without receiving a transplant. Motor behavior was assessed prior to the lesion as well as prior and 1, 3, and 5 weeks after the transplantations. At the end of the experimental period the number of graft-derived dopaminergic fibers growing into the host brain, the number of surviving dopaminergic neurons and graft volume were analyzed. In rats without a transplant, the density of dopaminergic fibers in the striatum was analyzed. We detected that NEP1-40 treatment significantly enhanced graft-derived dopaminergic fiber outgrowth as compared to controls while no effects were detected for graft volume and survival of grafted dopaminergic neurons. Notably, the enhanced dopaminergic fiber outgrowth was not sufficient to improve the functional recovery as compared to controls. Moreover, NEP1-40 infusions in hemi-parkinsonian rats without a transplant did not result in enhanced striatal dopaminergic fiber densities and consequently did not improve behavior. In sum, our findings demonstrate that antagonization of the Nogo-receptor 1 has the capacity to support the engraftment of transplanted mesencephalic tissue in an animal model of Parkinson's disease.

Simultaneous Transplantation of Fetal Ventral Mesencephalic Tissue and Encapsulated Genetically Modified Cells Releasing GDNF in a Hemi-Parkinsonian Rat Model of Parkinson's Disease.

Transplantation of fetal ventral mesencephalic (VM) neurons for Parkinson's disease (PD) is limited by poor survival and suboptimal integration of grafted tissue into the host brain. In a 6-hydroxydopamine rat model of PD, we investigated the feasibility of simultaneous transplantation of rat fetal VM tissue and polymer-encapsulated C2C12 myoblasts genetically modified to produce glial cell line-derived neurotrophic factor (GDNF) or mock-transfected myoblasts on graft function. Amphetamine-induced rotations were assessed prior to transplantation and 2, 4, 6 and 9 wk posttransplantation. We found that rats grafted with VM transplants and GDNF capsules showed a significant functional recovery 4 wk after implantation. In contrast, rats from the VM transplant and mock-capsule group did not improve at any time point analyzed. Moreover, we detected a significantly higher number of tyrosine hydroxylase immunoreactive (TH-ir) cells per graft (2-fold), a tendency for a larger graft volume and an overall higher TH-ir fiber outgrowth into the host brain (1.7-fold) in the group with VM transplants and GDNF capsules as compared to the VM transplant and mock-capsule group. Most prominent was the TH-ir fiber outgrowth toward the capsule (9-fold). Grafting of GDNF-pretreated VM transplants in combination with the implantation of GDNF capsules resulted in a tendency for a higher TH-ir fiber outgrowth into the host brain (1.7-fold) as compared to the group transplanted with untreated VM transplants and GDNF capsules. No differences between groups were observed for the number of surviving TH-ir neurons or graft volume. In conclusion, our findings demonstrate that simultaneous transplantation of fetal VM tissue and encapsulated GDNF-releasing cells is feasible and support the graft survival and function. Pretreatment of donor tissue with GDNF may offer a way to further improve cell transplantation approaches for PD.

GDNF family ligands display distinct action profiles on cultured GABAergic and serotonergic neurons of rat ventral mesencephalon.

Glial-cell-line-derived neurotrophic factor (GDNF), neurturin (NRTN), artemin (ARTN) and persephin (PSPN), known as the GDNF family ligands (GFLs), influence the development, survival and differentiation of cultured dopaminergic neurons from ventral mesencephalon (VM). Detailed knowledge about the effects of GFLs on other neuronal populations in the VM is essential for their potential application as therapeutic molecules for Parkinson's disease. Hence, in a comparative study, we investigated the effects of GFLs on cell densities and morphological differentiation of gamma-aminobutyric acid-immunoreactive (GABA-ir) and serotonin-ir (5-HT-ir) neurons in primary cultures of E14 rat VM. We observed that all GFLs [10 ng/ml] significantly increased GABA-ir cell densities (1.6-fold) as well as neurite length/neuron. However, only GDNF significantly increased the number of primary neurites/neuron, and none of the GFLs affected soma size of GABA-ir neurons. In contrast, only NRTN treatment significantly increased 5-HT-ir cells densities at 10 ng/ml (1.3-fold), while an augmentation was seen for GDNF and PSPN at 100 ng/ml (2.4-fold and 1.7-fold, respectively). ARTN had no effect on 5-HT-ir cell densities. Morphological analysis of 5-HT-ir neurons revealed a significant increase of soma size, number of primary neurites/neuron and neurite length/neuron after GDNF exposure, while PSPN only affected soma size, and NRTN and ARTN failed to exert any effect. In conclusion, we identified GFLs as effective neurotrophic factors for VM GABAergic and serotonergic neurons, demonstrating characteristic individual action profiles emphasizing their important and distinct roles during brain development.

Creatine supplementation improves neural progenitor cell survival in Huntington's disease.

Preclinical and clinical studies suggest that striatal transplantation of neural stem cells (NSCs) and neural progenitor cells (NPCs) may be an appealing and valuable system for treating Huntington's disease. Nevertheless, for a neural replacement to become an effective translational treatment for Huntington's disease, a certain number of difficulties must be addressed, including how to improve the integration of transplanted cell grafts with the host tissue, to elevate the survival rates of transplanted cells, and to ensure their directed differentiation into specific neuronal phenotypes. Research focusing on the translational applications of creatine (Cr) supplementation in NSC and NPC cell replacement therapies continues to offer promising results, pointing to Cr as a factor with the potential to improve cell graft survivability and encourage differentiation toward GABAergic phenotypes in models of striatal transplantation. Here, we evaluate research examining the outcomes of Cr supplementation and how the timing of supplementation regimes may affect their efficacy. The recent studies indicate that Cr's effects vary according to the developmental stage of the cells being treated, noting the dynamic differences in creatine kinase expression over the developmental stages of differentiating NPCs. This research continues to move Cr supplementation closer to the widespread clinical application and suggests such techniques warrant further examination.

Creatine supplementation improves dopaminergic cell survival and protects against MPP+ toxicity in an organotypic tissue culture system.

Cell replacement therapy using mesencephalic precursor cells is an experimental approach for the treatment of Parkinson's disease (PD). A significant problem associated with this procedure is the poor survival of grafted neurons. Impaired energy metabolism is considered to contribute to neuronal cell death after transplantation. Creatine is a substrate for mitochondrial and cytosolic creatine kinases (CK) and buffers cellular ATP resources. Furthermore, elevated cellular creatine levels facilitate metabolic channeling and show antiapoptotic properties. Exogenous creatine supplementation therefore might offer a tool for improvement of dopaminergic neuron survival. The present study aimed at investigating the effects of creatine on cell survival of rat embryonic day 14 (E14) ventral mesencephalic neurons grown as organotypic free-floating roller tube (FFRT) cultures. We found that the brain-specific isoform of CK (BB-CK) and the ubiquitous mitochondrial isoform (uMt-CK) are expressed at high levels in FFRT cultures and colocalize with tyrosine hydroxylase immunoreactive (TH-ir) cells. Exposure of these cultures to creatine induced an increase in the content of the BB-CK isotype. Creatine (5 mM) administration starting at day in vitro (DIV) 7 resulted in a significant increase (+35%) in TH-ir cell density at DIV21. In addition, we observed that creatine treatment provided neuroprotection against 1-methyl-4-phenyl pyridinium ion (MPP+)-induced TH-ir cell loss in the FFRT culture system, resulting in a significantly higher density (+19%) of TH-ir neurons in creatine-treated cultures compared to corresponding controls. The decrease of TH-ir neurons in the MPP+-treated group corresponded with an increase in immunoreactivity for active caspase-3, an effect that was not seen in the group receiving creatine supplementation. In conclusion, our data imply that creatine administration is beneficial for the survival of TH-ir neurons encountering harmful conditions.

Effect of GDNF on differentiation of cultured ventral mesencephalic dopaminergic and non-dopaminergic calretinin-expressing neurons.

Glial cell line-derived neurotrophic factor (GDNF) is a potent survival factor for ventral mesencephalic (VM) dopaminergic neurons. Subpopulations of dopaminergic and non-dopaminergic VM neurons express the calcium-binding proteins calbindin (CB) and calretinin (CR). Characterization of the actions of GDNF on distinct subpopulations of VM cells is of great importance for its potential use as a therapeutic molecule and for understanding its role in neuronal development. The present study investigated the effects of GDNF on the survival and morphological differentiation of dopaminergic and non-dopaminergic neurons in primary cultures of embryonic day (E) 18 rat VM. As expected from our results obtained using E14 VM cells, GDNF significantly increased the morphological complexity of E18 CB-immunoreractive (CB-ir), tyrosine hydroxylase (TH)-ir, and CR-ir neurons and also the densities of CB-ir and TH-ir neurons. Interestingly, densities of E18 CR-ir neurons, contrarily to our previous observations on E14 CR-ir neurons, were significantly higher after GDNF treatment (by 1.5-fold). Colocalization analyses demonstrated that GDNF increased the densitiy of dopaminergic neurons expressing CR (TH+/CR+/CB-), while no significant effects were observed for TH-/CR+/CB- cell densities. In contrast, we found that GDNF significantly increased the total fiber length (2-fold), number of primary neurites (1.4-fold), number of branching points (2.5-fold), and the size of neurite field per neuron (1.8-fold) of the non-dopaminergic CR-expressing neurons (TH-/CR+/CB-). These cells were identified as GABA-expressing neurons. In conclusion, our findings recognize GDNF as a potent differentiation factor for the development of VM dopaminergic and non-dopaminergic CR-expressing neurons.

The GDNF family members neurturin, artemin and persephin promote the morphological differentiation of cultured ventral mesencephalic dopaminergic neurons.

Neurturin (NRTN), artemin (ARTN), persephin (PSPN) and glial cell line-derived neurotrophic factor (GDNF) form a group of neurotrophic factors, also known as the GDNF family ligands (GFLs). They signal through a receptor complex composed of a high-affinity ligand binding subunit, postulated ligand specific, and a common membrane-bound tyrosine kinase RET. Recently, also NCAM has been identified as an alternative signaling receptor. GFLs have been reported to promote survival of cultured dopaminergic neurons. In addition, GDNF treatments have been shown to increase morphological differentiation of tyrosine hydroxylase immunoreactive (TH-ir) neurons. The present comparative study investigated the dose-dependent effects of GFLs on survival and morphological differentiation of TH-ir neurons in primary cultures of E14 rat ventral mesencephalon. Both NRTN and ARTN chronically administered for 5 days significantly increased survival and morphological differentiation of TH-ir cells at all doses investigated [0.1-100 ng/ml], whereas PSPN was found to be slightly less potent with effects on TH-ir cell numbers and morphology at 1.6-100 ng/ml and 6.3-100 ng/ml, respectively. In conclusion, our findings identify NRTN, ARTN and PSPN as potent neurotrophic factors that may play an important role in the structural development and plasticity of ventral mesencephalic dopaminergic neurons.

Rapid Recurrence of a Benign Meningial Perineurioma.

BACKGROUND: We describe a rare case of a rapidly recurring benign meningial-based perineurioma. Clinical, radiologic, and pathologic features of a rapidly recurring falxial perineurioma are described; the perineurioma was discovered incidentally in an 86-year-old woman. CASE DESCRIPTION: Due to progressive gait disturbances and radiologically proven progression after a 3-year symptom-free interval, subtotal resection of a large falxial-based meningeal tumor was performed. CONCLUSIONS: The pathologic examination confirmed the diagnosis of a perineurioma (World Health Organization grade I). Follow-up magnetic resonance tomography 5 months later due to neurologic deterioration revealed an abnormally rapidly growing and extensive local tumor recurrence. Due to the mass effect, reoperation was performed and adjuvant radiation of 20 Gy to the tumor bed was implemented thereafter. Meningeal-based perineuriomas of the central nervous system are extremely rare, and literature on proper management is scarce. Although histologic classification reveals a benign lesion, follow-up may be considered for this type of tumor.