Objective/Hypothesis: The role of fungal pathogens in the etiology of nasal polyposis remains unclear. The aim of this study was to determine whether there was a correlation between the presence of Alternaria-specific immunoglobulin (Ig)E antibodies, eosinophilic inflammation, and the development of nasal polyps.Study Design: Prospective study.Methods: Serum and nasal tissue homogenates from 21 patients with manifestations of chronic sinusitis with nasal polyps were compared with specimens from 13 chronic sinusitis patients without polyps and 8 healthy controls. The Phadia ImmunoCAP and enzyme-linked immunosorbent assay were used to quantify levels of total IgE and Alternaria-specific (IgE, IgG, and IgA) antibodies. Eosinophil cationic protein (ECP) and tryptase levels were measured in tissue homogenates, whereas the inflammatory response was evaluated using tissue eosinophil counts in tissue samples.Results: Serum analysis revealed no difference in the levels of total IgE and Alternaria-specific IgE, IgG, and IgA antibodies between the study groups. In contrast, the levels of Alternaria-specific IgE in tissue with polyps were significantly higher than in nonpolyp tissue. Increases in total tissue IgE paralleled increased levels of Alternaria-specific IgG and IgA antibodies in chronic sinusitis with nasal polyps as compared with control groups. A positive correlation was found between Alternaria-specific IgE and ECP in tissue. Increased mean levels of ECP corresponded to increased eosinophil counts in the group of patients with polyps.Conclusions: Alternaria-specific IgE and eosinophilic inflammation in nasal tissue correlates with the incidence of nasal polyps irrespective of specific IgE antibodies in serum. Together, the correlation between the local immune responses and the eosinophilic inflammation in nasal polyps suggests a possible role of Alternaria in the pathogenesis of nasal polyposis.

NMHC-IIa, a nonmuscle myosin heavy chain isoform encoded by MYH9, is expressed in sensory hair cells and its dysfunction is associated with syndromic and nonsyndromic hearing loss. In this study, we investigate the ultrastructural distribution of NMHC-IIa within murine hair cells to elucidate its potential role in hair cell function. Using previously characterized anti-mouse NMHC-IIa antibody detected with immunogold labelling, NMHC-IIa was observed in the stereocilia, in the cytosol along the plasma membrane, and within mitochondria. Within stereocilia, presence of NMHC-IIa is observed throughout its length along the actin core, from the center to the periphery and at its base in the cuticular plate, suggesting a potential role in structural support. Within the sensory hair cells, NMHC-IIa was distributed throughout the cytoplasm and along the plasma membrane. A novel finding of this study is the localization of NMHC-IIa within the mitochondria, with the majority of the label along its inner membrane folds. The presence of NMHC-IIa within heterogeneous areas of the hair cell suggests that it may play different functional roles in these distinct regions. Thus, mutant NMHC-IIa may cause hearing loss by affecting hair cell dysfunction through structural and or functional disruption of its stereocilia, plasma membrane, and/or mitochondria.

The purpose of this study is to determine the feasibility of the direct matrix-assisted laser desorption/ionization (MALDI) identification of proteins in fixed T47D breast cancer cells and murine brain tissues. The ability to identify proteins from cells and tissue may lead to biomarkers that effectively predict the onset of defined disease states, and their dynamic behavior could be an important hint for drug target discoveries. Direct tissue application of trypsin allows protein identification in cells and tissues, while maintaining spatial integrity and intracellular organization. Using a chemical printer, matrix was co-registered on trypsinized human T47D breast cancer cells and cryo-preserved sections of murine brain tissue, followed by MALDI post-source decay (PSD) or MALDI collision-induced dissociation (CID), respectively. Mass-to-charge (m/z) data from the cells and brain tissues were processed using Mascot software interrogation of the National Center for Biotechnology Information (NCBI) database. Histone H2B was identified from cultured T47D human breast cancer cells. Tubulin beta2 was identified from mouse brain cortex following an induced stroke. These results suggest that MALDI PSD/CID, combined with bioinformatics, can be used for the direct identification of proteins from cells and tissues. Refinements in preparation techniques may improve this approach to provide a tool for quantitative proteomics and clinical analysis.

Genome-encoded microRNAs (miRNAs) are potent regulators of gene expression. The significance of miRNAs in various biological processes has been suggested by studies showing an important role of these small RNAs in regulation of cell differentiation. However, the role of miRNAs in regulation of differentiated cell physiology is not well established. Mature neurons express a large number of distinct miRNAs, but the role of miRNAs in postmitotic neurons has not been examined. Here, we provide evidence for an essential role of miRNAs in survival of differentiated neurons. We show that conditional Purkinje cell-specific ablation of the key miRNA-generating enzyme Dicer leads to Purkinje cell death. Deficiency in Dicer is associated with progressive loss of miRNAs, followed by cerebellar degeneration and development of ataxia. The progressive neurodegeneration in the absence of Dicer raises the possibility of an involvement of miRNAs in neurodegenerative disorders.

Animals move about their environment by sensing their surroundings and making adjustments according to need. All animals take the force of gravity into account when the brain and spinal cord undertake the planning and execution of movements. To what extent must animals learn to factor in the force of gravity when making neural calculations about movement? Are animals born knowing how to respond to gravity, or must the young nervous system learn to enter gravity into the equation? To study this issue, young rats were reared in two different gravitational environments (the one-G of Earth and the microgravity of low Earth orbit) that necessitated two different types of motor operations (movements) for optimal behavior. We inquired whether those portions of the young nervous system involved in movement, the motor system, can adapt to different gravitational levels and, if so, the cellular basis for this phenomenon. We studied two groups of rats that had been raised for 16 days in microgravity (eight or 14 days old at launch) and compared their walking and righting (ability to go from upside down to upright) and brain structure to those of control rats that developed on Earth. Flight rats were easily distinguished from the age-matched ground control rats in terms of both motor function and central nervous system structure. Mature surface righting predominated in control rats on the day of landing (R+O), while immature righting predominated in the flight rats on landing day and 30 days after landing. Some of these changes appear to be permanent. Several conclusions can be drawn from these studies: (1) Many aspects of motor behavior are preprogrammed into the young nervous system. In addition, several aspects of motor behavior are acquired as a function of the interaction of the developing organism and the rearing environment; (2) Widespread neuroanatomical differences between one-G- and microgravity-reared rats indicate that there is a structural basis for the adaptation to the rearing environment. These observations provide support for the idea that an animal's motor system adapts for optimal function within the environment experienced during a critical period in early postnatal life.

We investigated the temporal expression of the neural cell adhesion molecule, neurotrimin, in the rat cerebellum and the brainstem from birth to adulthood using immunoreactive labeling. A wave of expression accompanied the development of projection pathways extending from brainstem nuclei (pons/inferior olive) through the cerebellar peduncles into the arbor vitae and disappeared with myelination by P14. Immuno-EM revealed expression of neurotrimin on the surface of unmyelinated axons but not on astrocytes or oligodendroglia. With the development of the molecular and internal granular layers, intense labeling occurred on the surface of parallel fiber bundles, granule cells and mossy fibers. With synaptogenesis, each excitatory junction was labeled by the immunoreaction. By P21, neurotrimin reactivity decreased on the surfaces of neuronal somata, dendrites and axons but remained at excitatory synaptic contact sites in both the molecular and granular layers. The spatial-temporal expression pattern of neurotrimin suggests that this adhesion molecule plays a role in axonal fasciculation of specific cerebellar systems and may also be involved in the formation of excitatory synapses and their stabilization into adulthood.

During myelinogenesis, we found an exceedingly strong, transient expression of the α1E gene for the R-type voltage-gated calcium channel in CNS white matter. This immunoreactivity appeared in glial cells along specific pathways of the brainstem, cerebellum, and telencephalon. The reactivity followed a wave that progressed from the brainstem at P5, to the cerebellar peduncles by P8, the arbor vitae by P14, and the granular layer by P17. The reactivity-peaked about 3–4 days later and decreased gradually to become negligible in all areas before adulthood. Ultrastructural analysis confirmed that α1E immunoreactivity was located in oligodendroglial somata, their projections, paranodal wraps and loose myelin sheaths. There was a distinct association of the channel protein reactivity on oligodendroglial membranes in contact with the axon. We propose that glial projections, contacting axons, sense axonal firing through small K+ currents and open the high voltage R-type calcium channels to signal myelination.

Qualitative and quantitative changes were found in the cerebellar circuitry of old as compared to young rats. The old group had a reduced number of synapses (at least 30%), however, there was an increase in the size of remaining synaptic components (13.5% for spine head volume, 66% for bouton volume, and 17% for the area of synaptic contact zones). Furthermore, there were pronounced morphological changes in the older group appearing as: 1) prominent lipofuscin bodies in Purkinje cell somata, 2) numerous myelinated fibers in the lower part of the molecular layer, 3) tortuous Purkinje cell dendrites in a thinned molecular layer, and 4) abundant vacuolar profiles and membrane swirls in small and intermediate-sized dendrites. Our findings suggest that Purkinje cell dendrites are dying-back reducing the target field for granule cells and that remaining synaptic sites compensate by increasing synaptic contact area as well as the size of pre- and postsynaptic structures.

This study was undertaken to illustrate the potential for subarachnoid injection during retrobulbar block as a cause of respiratory arrest. Cadaver orbits were used to document the connection between the optic nerve sheath and the subarachnoid space. Following dissections of the orbits on one side of 24 cadavers, the optic nerve sheaths were identified and injected with 0.5 ml of water for measurement of pressure generated during injection. This was followed by intrasheath injection of equal volume of methylene blue for demonstrating the subarachnoid space surrounding the optic nerves. All injections were performed with a 1-ml syringe with a one-and-one-half-inch 22-G needle over a period of 10 s. The blue dye was found to track along the subarachnoid space of the optic nerve sheath to the chiasmatic cistern in the middle cranial fossa. Retrobulbar injections were performed on the contralateral undissected orbits and intrascleral injections were performed on undissected eyes. The size of the syringes, the gauge of the needles, and the speed of injection were uniform for all injections. The pressure generated by injection into the optic nerve sheath or intrascleral injection (approximately 138 mmHg) was three- to fourfold that produced by injection into the retrobulbar adipose tissue (approximately 35 mmHg) (P less than 0.05). The authors conclude that any resistance encountered during retrobulbar block should serve as a warning signal, mandating redirection of the needle, in order to prevent subarachnoid injection.

Our previous investigations demonstrated an increase in the size of remaining synaptic sites as an intermediate or possible alternative to sprouting plasticity. The total amount of postsynaptic contact area remained relatively constant for each target neuron even though there was a marked decrease in the number of sites on these neurons. In addition, enlarged boutons containing numerous synaptic vesicles were positioned adjacent to enlarged postsynaptic sites.

Chronic neurological deficits have been described in patients after presumed accidental subarachnoid injection of 2-chloroprocaine-CE (Nesacaine-CE; N-CE) intended for epidural block. This study investigated the possible role of pure 2-chloroprocaine (2-CP) and sodium bisulfite, two components of Nesacaine-CE, in causing these complications when injected separately into the lumbar subarachnoid space of neurologically intact awake rabbits. Repeated 2–4-mg spinal anesthetic doses of pure 2-CP in lactated Ringer's solution did not produce chronic hindlimb paralysis even though accumulated doses reached 50 mg. However, 1.2–2.4 mg of sodium bisulfite, the antioxidant in N-CE added to prolong shelf-life, resulted in irreversible hindlimb paralysis in 12 out of 14 animals. This amount of bisulfite is contained in 12–24 mg of 2% N-CE. The demonstration that persistent paralysis resulted from low dosages of sodium bisulfite contained in commercially available 2-CP requires revaluation of the suitability of this antioxidant for products prepared for intrathecal use.

Mechanosensitive cilia are vital to signaling and development across many species. In sensory hair cells, sound and movement are transduced by apical hair bundles. Each bundle is comprised of a single primary cilium (kinocilium) flanked by multiple rows of actin-filled projections (stereocilia). Extracellular tip links that interconnect stereocilia are thought to gate mechanosensitive channels. In contrast to stereocilia, kinocilia are not critical for hair-cell mechanotransduction. However, by sequentially imaging the structure of hair bundles and mechanosensitivity of individual lateral-line hair cells in vivo, we uncovered a central role for kinocilia in mechanosensation during development. Our data demonstrate that nascent hair cells require kinocilia and kinocilial links for mechanosensitivity. Although nascent hair bundles have correct planar polarity, the polarity of their responses to mechanical stimuli is initially reversed. Later in development, a switch to correctly polarized mechanosensitivity coincides with the formation of tip links and the onset of tip-link-dependent mechanotransduction.

This chapter talks about the primate cerebellar cortex. The discussion is interspersed in the presentation to explain immediately the interpretations given the electron micrographs. The capricious method of Golgi, which now and then reveals the external morphology of a nerve cell and all its processes more completely than any other technique, has been most successful when applied to the cerebellar cortex. In the present study features of the cerebellar cortex are illustrated with Golgi impregnations of the macaque monkey (Macaca mulatta) and with electron micrographs of man, the macaque monkey (Macaca mulatta) and the squirrel monkey (Suimiri sciureus).Olivocerebellar fibers, conceivably, drive the intracerebellar nuclei and continuing on as climbing fibers stimulate the Purkinje cells. The climbing fiber-Purkinje cell ratio may be one to one. Purkinje cells actively inhibit the cerebellar nuclei and Deiters nucleus. This inhibition is continuously modified by circuits activated by mossy fibers, arriving from spinal, reticular, vestibular, pontine and tegmental sources.