Faculty Bibliography

INTRODUCTION/BACKGROUND:Primary cervical and oromandibular dystonia (CD and OMD, respectively) are well-recognized movement disorders, often treated with botulinum toxin (BTx). In contrast, dystonia related to acute brain injuries is not well delineated. Our objective was to define in neurocritically ill patients the clinical characteristics of CD and OMD and to investigate the safety of BTx. METHODS:All acutely brain-injured patients admitted to a neurocritical care unit over a 10-month period were prospectively screened for CD and OMD. Clinical characteristics, etiology of brain injury, and pattern of dystonia were analyzed. Patients with clinically significant CD and OMD were treated with BTx and followed for 12 weeks. RESULTS:Of 165 patients screened, 33 had new-onset CD or OMD. Of 21 patients enrolled, 14 had CD, 5 had OMD, and 2 had both. The pattern of brain injury included 13 cerebral hemorrhages, 6 ischemic strokes, 1 status epilepticus, and 1 unclear etiology. Improvement after BTx was seen in four of seven patients with CD and two of four with OMD; no adverse effects occurred. Spontaneous improvement was recorded in 7 of 11 nontreated patients with CD or OMD. CONCLUSIONS:Acute secondary CD or OMD, associated with a variety of causes, was identified in 20% of acutely brain-injured patients. The temporal profile of dystonia onset and resolution in these patients was variable. Treatment with BTx in the neurocritical care setting seems to be safe. Future, larger scale randomized studies should evaluate the effectiveness of BTx treatment in this patient population.

Chronic mitochondrial dysfunction, in particular of complex I, has been strongly implicated in the dopaminergic neurodegeneration in Parkinson's disease. To elucidate the mechanisms of chronic complex I disruption-induced neurodegeneration, we induced differentiation of immortalized midbrain dopaminergic (MN9D) and non-dopaminergic (MN9X) neuronal cells, to maintain them in culture without significant cell proliferation and compared their survivals following chronic exposure to nanomolar rotenone, an irreversible complex I inhibitor. Rotenone killed more dopaminergic MN9D cells than non-dopaminergic MN9X cells. Oxidative stress played an important role in rotenone-induced neurodegeneration of MN9X cells, but not MN9D cells: rotenone oxidatively modified proteins more in MN9X cells than in MN9D cells and antioxidants decreased rotenone toxicity only in MN9X cells. MN9X cells were also more sensitive to exogenous oxidants than MN9D cells. In contrast, disruption of bioenergetics played a more important role in MN9D cells: rotenone decreased mitochondrial membrane protential and ATP levels in MN9D cells more than in MN9X cells. Supplementation of cellular energy with a ketone body, D-beta-hydroxybutyrate, decreased rotenone toxicity in MN9D cells, but not in MN9X cells. MN9D cells were also more susceptible to disruption of oxidative phosphorylation or glycolysis than MN9X cells. These findings indicate that, during chronic rotenone exposure, MN9D cells die primarily through mitochondrial energy disruption, whereas MN9X cells die primarily via oxidative stress. Thus, intrinsic properties of individual cell types play important roles in determining the predominant mechanism of complex I inhibition-induced neurodegeneration.

OBJECTIVE:Excitotoxicity is a multistep process that results in either necrosis or apoptosis. It has been associated with neuronal death in trauma, ischemia, and neurodegeneration. The final step in necrotic cell death is the rupture of a cell's plasma membrane; repair of this membrane rupture is a potentially powerful technique of neuroprotection. Poloxamer 188 (P-188) is a synthetic surfactant that seals experimentally porated membranes. This study investigated the usefulness and time dependence of intrathecal P-188 in protecting neurons in an in vivo model of excitotoxicity in the rat. METHODS:Twenty-eight Sprague-Dawley rats underwent striatal infusion of quinolinic acid to produce a spherical excitotoxic lesion. Each animal then received either vehicle or P-188 at 10 minutes, 4 hours, or both time points after surgery by direct cisterna magna injection. Animals were killed at 1 week, and brains were stained immunohistochemically for the neuronal marker Neu-N. Volumes of neuronal loss were calculated and compared between groups by analysis of variance. RESULTS:All animals were found to have spherical, stereotyped lesions. The animals that received intrathecal poloxamer at the early injection time had statistically smaller lesions (8.16 +/- 6.12 mm(3); n = 5; P = 0.0015) than controls (18.25 +/- 11.42 mm(3); n = 11). Those animals that received poloxamer at both injection times also had statistically smaller lesions (10.57 +/- 9.00 mm(3); n = 7; P = 0.0095). The group that received poloxamer at the late injection time only did not have significantly decreased lesion size (14.86 +/- 7.95 mm(3); n = 5). CONCLUSION/CONCLUSIONS:Intrathecal P-188 reduces neuronal loss after excitotoxic injury in the rat only when delivered immediately after the toxin. This observation confirms the potential of surfactant molecules as neuroprotectants but predicts that their usefulness is best realized by early and potentially ongoing treatment.

OBJECT/OBJECTIVE:The surfactant, poloxamer 188 (P-188), has been found to protect against tissue injury in various experimental models. Its protective mechanism may involve the effects of the surfactant against oxidative stress and inflammation. The authors investigated the role of P- 188 in the reduction of tissue injury and macrophage response in a model of excitotoxic brain injury in the rat striatum. METHODS:Fifteen Sprague-Dawley rats underwent stereotactic injection of 120 nmol of quinolinic acid into the striatum and received intracisternal injection of vehicle or P-188 (40 mg/kg) at 10 minutes and 4 hours postinjury. Rats were killed after 1 week, and the histological score was determined based on the degree of overall tissue injury (Grades 1-4) at the lesion site. The number of macrophages within the lesioned striatum was compared with that found within the striatum on the nonoperated contralateral side. The scores related to tissue damage and the macrophage ratios of each group were then compared using t-tests. Striatal injection of the toxin produced a lesion characterized by necrosis and inflammation surrounding the injection site in all six control animals. In rats in which intracisternal P-188 was administered, significantly less tissue injury was demonstrated (mean score 2.45 +/- 0.74) than in controls (mean score 3.14 +/- 0.75) (p = 0.045). The rats that received intracisternal surfactant also had significantly less macrophage infiltrate (mean ratio 1.06 +/- 0.18) than control animals (mean ratio 2.00 +/- 0.48) (p = 0.004). CONCLUSIONS:The surfactant P-188 reduces tissue loss and macrophage infiltrate after excitotoxic brain injury in the rat. Possible mechanisms of this effect may include direct surfactant modulation of inflammatory cell membrane fluidity.

Membrane repair of damaged neurons by surfactant poloxamers has been noted in experimental spinal cord injury and in vitro excitotoxicity. We examined poloxamer-188 (P-188)-mediated neuroprotection in a rat model of glutamate toxicity. Quinolinate was infused into the striatum followed 10 min and 4 h later by P-188 administered either i.v. or intracisternally (i.c.), or by vehicle. Mean neuronal loss examined volumetrically 7 days later in control animals was 50% greater (P < 0.01) than after i.c. P-188 treatment; control lesion volumes were 38% greater than lesion volumes after i.v. P-188 treatment; however, that comparison did not reach significance. This robust protection against glutamate toxicity may predict P-188-mediated neuroprotection against a broad range of clinically relevant neural insults.

Chronic L-3,4-dihydroxyphenylalanine (L-DOPA) therapy in Parkinson's disease (PD) is complicated by motor response fluctuations and dyskinesia. The relative contributions of disease severity and chronic L-DOPA therapy to the development of motor fluctuation are not well defined clinically. Experimental studies have been limited partly because models for the antiparkinsonian effects on akinesia have not been employed. Therefore, we employed a model of akinesia using forepaw adjusting steps that have been well characterized to reflect the effect of lesions and the antiparkinsonian effect of dopaminergic drugs and transplants. We administered L-DOPA (12.5 mg/kg) intermittently for 4 weeks to rats with severe nigrostriatal lesions produced by injecting 6-hydroxydopamine into the medial forebrain bundle. The peak magnitude responses to L-DOPA increased after treatment compared to the pretreatment baseline. The latency to peak response to L-DOPA became shorter and reversed after the discontinuation of treatment. The duration of response showed minor changes. The pattern of changes in response to apomorphine was similar to that of L-DOPA except that the peak magnitude did not increase despite chronic L-DOPA treatment. The changes in D1 and D2 receptor binding did not correlate with behavioral changes. In summary, long-term intermittent L-DOPA treatment resulted in priming of antiparkinsonian effects on improving akinesia in a rat model of severe PD. These observed changes do not mirror all aspects of motor response fluctuations in advanced PD patients and suggest differential contributions of dopaminergic treatment and lesion severity to motor complication patterns.

Dopaminergic (DA) neurons in the ventral midbrain nuclei, substantia nigra pars compacta (SNc, A9) and ventral tegmental area (VTA, A10), play important roles in the control of movement, emotion, cognition, and reward related behavior. Although several transcription factors have been shown to be critical for midbrain DA neuron development, there has been no report of factor(s) that differentially regulate individual DA neuronal groups. Based on its highly restricted expression in the SNc and VTA in the brain, we hypothesize that the homeobox transcription factor Pitx3 may critically regulate the development of ventral midbrain DA neurons. In this study, we report that in Pitx3-deficient ak/ak mice, DA neurons in the SNc and the nigrostriatal pathway fail to develop properly, and DA levels are reduced to 10% of the wild type mice in the dorsal striatum. On the contrary, A10 neurons are intact in ak/ak mice and DA levels within their projection areas are not affected. This region-specific defect was already evident in newborn mice, suggesting that the defect had occurred during the early stages of mouse development. Taken together, our results indicate that Pitx3 is the first known transcription factor that may critically and selectively control proper development of A9 DA neurons and the nigrostriatal pathway. This observation is of great importance in understanding the mechanisms of DA neuron development and may also help us to understand the mechanism of selective degeneration of A9 DA neurons in Parkinson's disease and to devise novel therapeutic approaches for the disorder.

Gene therapy methods have continued to develop rapidly, and many initial limitations that hampered clinical application have been overcome. Thus serious consideration of clinical application of gene therapy is warranted for selected disorders in which the pathogenesis is well defined. Parkinson's disease has been the most extensively studied target of gene therapy for central nervous system disorders and shares many features with pediatric neurotransmitter diseases. Neurotransmitter replacement therapy using catecholamine-synthesizing genes and delivery of neurotrophic factors such as glial cell line-derived neurotrophic factors has been successful in animal models of Parkinson's disease. Application of gene therapy for pediatric neurotransmitter diseases will require delineating the optimal set of genes to correct the consequences of the deficiencies. The optimal anatomical targets and proper timing of the gene replacement must be understood. Safety of gene therapy vehicles and the ability to regulate gene expression will be essential for eventual clinical application.