Different routes of heroin intake cause various heroin-induced leukoencephalopathies.

OBJECTIVE: Toxic leukoencephalopathy is a rare but critical neurological disorder in heroin abusers. Our aim is to compare the clinical manifestations, brain MRIs and prognoses of heroin-induced leukoencephalopathy by different intake routes. METHODS: We present two patients with toxic leukoencephalopathy caused by intravenous (IV) injection of heroin and 48 additional cases from systematic reviews of the literature published between 1994 and 2018. RESULTS: Among the 50 heroin abusers who developed leukoencephalopathy, inhalation was the most popular route (60%), followed by IV injection (30%) and snorting (10%). Mental changes, mutism and urine/fecal incontinence were the major symptoms in patients who IV injected heroin, while cerebellar ataxia and dysarthria were more common among those who inhaled heroin. Delayed-onset encephalopathy uniquely occurred in those who IV injected heroin, whereas progressive encephalopathy was more commonly observed in those who inhaled heroin. Clinical improvement was observed in 60% of patients, the overall mortality rate was 12%, and higher mortality was observed in patients who used the inhalation route (16.7%). The hallmarks on the MRIs of those who inhaled heroin were posterior to anterior involvement of the cerebral white matter and lesions in the posterior limbs of the internal capsules, cerebellum and brainstem. In contrast, those who IV injected heroin had more frequent lesions in the subcortical U fibers and the genu of the internal capsules. CONCLUSION: These data could help physicians make an early diagnosis and predict prognosis and suggest that prompt antioxidative or symptomatic treatments might reduce the long-term consequences and mortality of heroin-induced leukoencephalopathy.

High-Frequency Stimulation of the Subthalamic Nucleus Activates Motor Cortex Pyramidal Tract Neurons by a Process Involving Local Glutamate, GABA and Dopamine Receptors in Hemi-Parkinsonian Rats.

Deep brain stimulation (DBS) is widely used to treat advanced Parkinson's disease (PD). Here, we investigated how DBS applied on the subthalamic nucleus (STN) influenced the neural activity in the motor cortex. Rats, which had the midbrain dopaminergic neurons partially depleted unilaterally, called the hemi-Parkinsonian rats, were used as a study model. c-Fos expression in the neurons was used as an indicator of neural activity. Application of high-frequency stimulation (HFS) upon the STN was used to mimic the DBS treatment. The motor cortices in the two hemispheres of hemi-Parkinsonian rats were found to contain unequal densities of c-Fos-positive (Fos+) cells, and STN-HFS rectified this bilateral imbalance. In addition, STN-HFS led to the intense c-Fos expression in a group of motor cortical neurons which exhibited biochemical and anatomical characteristics resembling those of the pyramidal tract (PT) neurons sending efferent projections to the STN. The number of PT neurons expressing high levels of c-Fos was significantly reduced by local application of the antagonists of non-N-methyl-D-aspartate (non-NMDA) glutamate receptors, gammaaminobutyric acid A (GABAA) receptors and dopamine receptors in the upper layers of the motor cortex. The results indicate that the coincident activations of synapses and dopamine receptors in the motor cortex during STN-HFS trigger the intense expression of c-Fos of the PT neurons. The implications of the results on the cellular mechanism underlying the therapeutic effects of STN-DBS on the movement disorders of PD are also discussed.

In vitro growth conditions and development affect differential distributions of RNA in axonal growth cones and shafts of cultured rat hippocampal neurons.

Local synthesis of proteins in the axons participates in axonogenesis and axon guidance to establish appropriate synaptic connections and confer plasticity. To study the transcripts present in the growth cones and axonal shafts of cultured rat hippocampal neurons, two chip devices, differing in their abilities to support axonal growth and branching, are designed and employed here to isolate large quantities of axonal materials. Cone-, shaft- and axon-residing transcripts with amounts higher than that of a somatodendritic transcript, Actg1 (γ-actin), are selected and classified. Since the chips are optically transparent, distribution of transcripts over axons can be studied by fluorescence in situ hybridization. Three transcripts, Cadm1 (cell adhesion molecule 1), Nefl (neurofilament light polypeptide), and Cfl1 (non-muscle cofilin) are confirmed to be preferentially localized to the growth cones, while Pfn2 (profilin2) is preferentially localized to the shafts of those axons growing on the chip that restricts axonal growth. The different growing conditions of axons on chips and on conventional coverslips do not affect the cone-preferred localization of Cadm1 and shaft-preferred localization of Pfn2, but affect the distributions of Nefl and Cfl1 over the axons at 14th day in vitro. Furthermore, the distributions of Cadm1 and Nefl over the axons growing on conventional coverslips undergo changes during in vitro development. Our results suggest a dynamic nature of the mechanisms regulating the distributions of transcripts in axonal substructures in a manner dependent upon both growth conditions and neuronal maturation.

Cell-based analysis of Chikungunya virus membrane fusion using baculovirus-expression vectors.

Chikungunya virus infection has emerged in many countries over the past decade. There are no effective drugs for controlling the disease. To develop cell-based system for screening anti-virus drugs, a bi-cistronic baculovirus expression system was utilized to co-express viral structural proteins C (capsid), E2 and E1 and the enhanced green fluorescence protein (EGFP) in Spodoptera frugiperda insect cells (Sf21). The EGFP-positive Sf21 cells fused with each other and with uninfected cells to form a syncytium, allowing characterization of cholesterol and low pH requirements for syncytium formation. Western blot analysis showed three structural proteins were expressed in baculovirus infected cells. The structural proteins of Chikungunya virus that is required for cell fusion was determined with various recombinant baculoviruses bearing different lengths of the viral structural protein genes. Protein E1 was required for cell fusion and indicating that Chikungunya viral membrane fusion was a class II membrane fusion. It was also demonstrated that the heterologous expression of alphavirus monomeric E1 can induce insect cell fusions. Furthermore, this cell-based system provides a model for studying class II viral membrane fusion.

Cold-induced exodus of postsynaptic proteins from dendritic spines.

Dendritic spines are small protrusions on neuronal dendrites and the major target of the excitatory inputs in mammalian brains. Cultured neurons and brain slices are important tools in studying the biochemical and cellular properties of dendritic spines. During the processes of immunocytochemical studies of neurons and the preparation of brain slices, neurons were often kept at temperatures lower than 37 degrees C for varied lengths of time. This study sought to investigate whether and how cold treatment would affect the protein composition of dendritic spines. The results indicated that upon cold treatment four postsynaptic proteins, namely, alpha,beta-tubulins, calcium, calmodulin-dependent protein kinase IIalpha, and cytoplasmic dynein heavy chain and microtubule-associated protein 2, but not PSD-95 or AMPA receptors, exited from the majority of dendritic spines of cultured rat hippocampal neurons in a Gd(3+)-sensitive manner. The cold-induced exit of tubulins from dendritic spines was further found to be an energy-dependent process involving the activation of Gd(3+)-sensitive calcium channels and ryanodine receptors. The results thus indicate that changes in temperature, calcium concentration, and energy supply of the medium surrounding neurons would affect the protein composition of the dendritic spines and conceivably the protein composition of the subcellular organizations, such as the postsynaptic density, in the cytoplasm of dendritic spines.

Heavy chain of cytoplasmic dynein is a major component of the postsynaptic density fraction.

A protein with an apparent molecular size of 490 kDa was found in the postsynaptic density (PSD) fraction isolated from porcine cerebral cortices and rat forebrains, and this 490 kDa protein accounted for approximately 3% of the total protein of these samples. Matrix-assisted laser desorption ionization-time of flight mass spectrometric and Western blotting analyses consistently indicated that this 490 kDa protein consisted primarily of the heavy chain of cytoplasmic dynein (cDHC). Immunocytochemical analyses showed that cDHC was found in 92% and 89% of the phalloidin-positive protrusions that were themselves associated with discrete clusters of synaptophysin, a presynaptic terminal marker, and PSD-95, a postsynaptic marker, on neuronal processes, respectively. Quantitative Western blotting analyses of various subcellular fractions isolated from porcine cerebral cortices and rat forebrains further showed that not only the heavy but also the intermediate chains of dynein are enriched in the PSD fraction. Cytoplasmic dynein is a microtubule-associated motor protein complex that drives the movement of various cargos toward the minus ends of microtubules and plays many other diverse functions in the cell. Our results that cDHC is a major component of the PSD fraction, that both dynein heavy and intermediate chains are enriched in the PSD fraction and that cDHC is present in dendritic spines raise the possibilities that cytoplasmic dynein may play structural and functional roles in the postsynaptic terminal.