Neuroscience
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Comparative Study
Expression of TWIK-related acid sensitive K+ channels in capsaicin sensitive and insensitive cells of rat dorsal root ganglia.
Previous reports have demonstrated that small- to medium-diameter dorsal root ganglia (DRG) cells in rats can be subgrouped into individual cell types by patterns of voltage-activated currents. These cell types have consistent responses to algesic compounds and maintain characteristic histochemical phenotypes. Using immunocytochemical methods, we have now examined expression of TWIK (tandem of P domains in a weak inwardly rectifying K+ channel)-related acid sensitive K+ (TASK) channels, TASK-1, TASK-2 and TASK-3, in nine electrophysiologically identified small- to medium-diameter DRG cell types. ⋯ The co-expression of TASK-1 and TASK-3 in cell types 1, 4 and 6 suggests that these sensory afferents might contain functional heterodimeric channels. In peripheral sensory afferents, TASK channels have been implicated in the pain sensory transduction pathway, and can be modulated by anesthetics and neuroprotective agents. This study seeks to identify TASK channel populations in electrophysiologically characterized populations of putative nociceptive afferents.
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The study was aimed at investigating the expression and the activity of neuronal nitric oxide synthase, and of soluble guanylyl cyclase and phosphodiesterase activities that regulate guanosine 3',5'-cyclic monophosphate level in the midbrain, in a mouse model of PD using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine injections. Adult male mice of the C57/BL strain were given three i.p. injections of physiological saline or three i.p. injections of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine solution in physiological saline at 2 h intervals (summary 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine dose: 40 mg/kg), and were killed 3, 7, or 14 days later. mRNA, protein level, and/or activities of neuronal nitric oxide synthase, soluble guanylyl cyclase, phosphodiesterase and guanosine 3',5'-cyclic monophosphate were determined. Immunohistochemistry showed about 75% decrease in the number of tyrosine hydroxylase-positive neurons in the substantia nigra pars compacta. ⋯ The increases in guanylyl cyclase activity were found to occur exclusively due to increased maximal enzyme activity. No 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced change in phosphodiesterase activity has been detected in any brain region studied. 7-Nitroindazole prevented a significant increase in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced midbrain guanosine 3',5'-cyclic monophosphate level and neurodegeneration of dopaminergic neurons. These results raise the possibility that the nitric oxide/guanylyl cyclase/guanosine 3',5'-cyclic monophosphate signaling pathway may play a role in maintaining dopaminergic neurons function in substantia nigra pars compacta.
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Aiming to gain insights into the mechanisms of neuronal differentiation, we describe the first differential expression profiles of purified homogenous neural precursors (CD133+ cells from human fetal brain) with those of differentiated neurons from human fetal brain. The purity of the two populations of cells was verified by flow cytometry and immunocytochemistry, and cells were then processed for DNA microarray analysis. ⋯ In addition, we identified, and confirmed by reverse transcription-polymerase chain reaction and in situ hybridization, significant differential expression of platelet-derived growth factor receptor-alpha and insulin-like growth factor binding protein 4, indicating these factors as potential pro-neuronal differentiation factors. In summary, by using the microarray technique to perform a comparative analysis of the genes involved in the differentiation of neural precursors, enriched from the human fetus, we have identified hitherto unknown candidate genes and related signaling pathways that might play an essential role in neuronal differentiation.
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Comparative Study
Dendritic morphogenesis of cerebellar Purkinje cells through extension and retraction revealed by long-term tracking of living cells in vitro.
Cerebellar Purkinje cells have the most elaborate dendritic trees among the neurons in the CNS. To investigate the dynamic aspects of dendritic morphogenesis of Purkinje cells, we performed a long-term analysis of living cells in cerebellar cell cultures derived from glutamate decarboxylase 67-green fluorescent protein mice. Most Purkinje cells had several primary dendrites during the 25-day culture period. ⋯ Furthermore, treatment with an inhibitor of calcium/calmodulin-dependent protein kinase II reduced the number of primary dendrites specifically during 5-15 days in vitro, the culture period when the extension and retraction of primary dendrites occurred actively. Blockade of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid/kainate-type glutamate receptors also reduced the number of primary dendrites during the same culture period, while inhibition of glutamate transporters increased the number. These findings suggest that the final morphology of Purkinje cells is achieved not only through extension, but also through retraction of their dendrites, and that calcium/calmodulin-dependent protein kinase II and neuronal activity are involved in this dendritic morphogenesis.
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Comparative Study
C fragment of tetanus toxin hybrid proteins evaluated for muscle-specific transsynaptic mapping of spinal motor circuitry in the newborn mouse.
We investigated whether the non-toxic C fragment of tetanus toxin (TTC) fused to either beta-galactosidase or green fluorescent protein could be utilized to transsynaptically trace muscle-specific spinal circuitry in the neonatal mouse after i.m. injection into a single hindlimb muscle. We found that even with careful low volume injection (0.2-1.0 microl) into a single muscle (medial gastrocnemius), the TTC hybrid proteins spread rapidly to many other hindlimb muscles and to trunk musculature such that retrograde labeling of motoneurons could not be constrained to a single motoneuron pool. Retrogradely labeled motoneurons in the lower lumbar segments harboring the medial gastrocnemius motoneuron pool were first observed two hours after the medial gastrocnemius injection. ⋯ TTC injection procedure described here therefore provides an important tool for the study of presynaptic terminals onto motoneurons. However, additional technical modifications will be required to utilize TTC tracers for transsynaptic mapping of muscle-specific spinal motor circuitry in the neonatal mouse. We provide here a set of criteria for assessing the i.m. delivery of TTC tracers as a basis for future improvements in this technique.